Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 04/24/2026 has been entered.
DETAILED ACTION
Claims 2-3, 8, 10, 25-26 and 42 are cancelled. Claims 1, 4-7, 9, 11-24, 27-41 and 43-44 are pending. Claims 15-16, 20-24, 30-36, and 40-41 are withdrawn as being drawn to nonelected inventions. Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are under consideration.
Priority
The claims are not entitled to the priority date of foreign application EP18205852.9, because the foreign application does not provide adequate support or enablement for the following limitations recited in claim 1 and required by the dependent claims: “the C-terminal amino acid of the cellulase component is changed from alanine to proline (A228P)” (lines 3-4), and “the fusion protein has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14” (lines 13-15). MPEP 216 states that the foreign application may be considered in the same manner as if it had been filed in this country on the same date that it was filed in the foreign country.
Accordingly, claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are entitled to an effective filing date of 11/12/2019.
Claim Rejections - 35 USC § 112(a)
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention. The amendment filed on 04/24/2026 has introduced new matter into the claims.
Claim 1, as filed on 04/24/2026 recites a fusion protein comprising a cellulase component, a linker component, and a carbohydrate binding component, wherein: the cellulase component is an endoglucanase, wherein the C-terminal amino acid of the cellulase component is changed from alanine to proline (A228P); the linker component has at least 94% sequence identity with SEQ ID NO: 1, 2 or 3, and, according to the numbering corresponding to the SEQ ID NO 1, comprises the following characteristics: the amino acid at position 3 is S; the amino acid at position 8 is R; the amino acid at position 13 is S; and the amino acid at position 18 is S; and the carbohydrate binding component has capability to bind cellulose; wherein the fusion protein has a depilling and/or antipilling effect on fabric containing cellulose or cellulose derivative; and wherein the fusion protein has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14, in a liquid detergent in the presence of a protease.
Applicant asserts that no new matter was added in the amendment. See the remarks filed 04/24/2026 p. 10 paragraph 1. Applicant has not pointed out where the amended claims are supported, nor does there appear to be a written description of the underlined claim limitations above in the application as filed.
Claim 1 (and dependent claims) contains new matter because of the limitation requiring the C-terminal amino acid of the cellulase component to be changed from alanine to proline (A228P); and because of the limitation requiring the fusion protein to have at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14.
The specification and the original claims, as filed 05/04/2021, do not provide support for the full breadth encompassed by the limitation that requires the C-terminal amino acid of the cellulase component to be changed from alanine to proline (A228P). The specification states that: in an embodiment the C-terminal amino acid of the cellulase component is Pro, corresponding to the substitution A228P in SEQ ID NO: 14. See p. 22 lines 13-14. SEQ ID NO: 14 is the full length amino acid sequence of MA79+CBM (i.e. MA79 cellulase component + carbohydrate binding component) deriving from Melanocarpus albomyces 20K-cellulase including amino acids Met1 to Leu304. See p. 6 lines 26-28. The specification discloses that SEQ ID NO: 12 is the MA79 cellulase amino acid sequence from Met1 to Ala235. See p. 6 lines 21-23. Additional cellulase components disclosed include SEQ ID NOs: 4 (p. 6 line1), 6 (p. 6 line 6), 8 (p. 6 line 11), 10 (p. 6 line 16). Shown below is an alignment between the cellulases amino acid sequence disclosed (SEQ ID Nos: 4, 6, 8, 10 and 12) and the fusion protein SEQ ID NO: 14, which includes a truncated SEQ ID NO: 12 cellulase component with an A228P substitution. The C-terminal residues of the cellulases are boxed, and the A228P substitution is circled. Since claim 1 does not limit the fusion protein to SEQ ID NO: 14, there is no support commensurate in scope with the instantly claimed alanine to proline change.
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The specification and the original claims, as filed 05/04/2021, do not provide support for the full breadth encompassed by the claimed fusion protein that has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14. The specification teaches that linkers that have at least 90% sequence identity with SEQ ID NO: 1 have stability in liquid detergent containing proteases. See p. 22 lines 14-19. In example 3, the specification teaches testing the stability of fusion cellulases MA79+CBM (i.e. SEQ ID NO: 14) and MA79+CBML (i.e. SEQ ID NO: 19) by application tests as color revival using parental MA79 (i.e. SEQ ID NO: 12) for comparison. See p. 21 lines 6-8. The stability of fusion cellulases MA79+CBM and MA79+CBML is shown in figure 4. See p. 32 lines 3-4. The specification discloses that the stability of MA79+CBML measured as application performance was about 5 times better than that with MA79+CBM. See p. 32 line 7-9. The specification does not provide support commensurate in scope with the instant claims because the instant claims do not limit the fusion protein to the MA79+CBML (SEQ ID NO: 19) embodiment disclosed in example 3, nor does the specification provide support for a fusion protein that has at least a 5-fold increase in improved protease stability.
Such limitations recited in the instant claim 1 (and required by dependent claims), which did not appear in the specification or original claims, as filed, introduce new concepts and violate the description requirement of the first paragraph of 35 U.S.C 112. Applicant is required to provide sufficient written support for the limitations recited in the instant claims. Applicant can remove the new matter limitations from the claims to obviate this rejection.
Claim Rejections - 35 USC § 112(b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 1 recites the parenthetical phrase "(A228P)", renders the claim indefinite because it is unclear whether the limitation within the parentheses is part of the claimed invention. See MPEP § 2173.05(d). It is unclear whether claim 1 requires the C-terminal amino acid of the cellulase component to be changed from alanine to proline specifically at residue position 228 because the (A228P) limitation is recited as a parenthetical phrase. If the claim does require an A228P change, it is further unclear which position 228 is being referenced because the claim does not limit the sequence of the cellulase component.
Claims 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 depend from claim 1 and are rejected for the reason set forth above.
Claims 5 and 6 recite “the cellulase component has at least 80% sequence identity”, which render the claims indefinite because it is unclear whether the “at least 80% sequence identity” limitations include or exclude the C-terminal amino acid change from alanine to proline required in instant claim 1. Claim 5 requires the cellulase component to have at least 80% sequence identity over residues 18-224 with the SEQ ID NO: 4. It is unclear whether claim 5 is requiring the C-terminal residue to be position 224 of SEQ ID NO: 4, which is a proline residue. If the C-terminal residue is not required to be position 224 of SEQ ID NO: 4, then it is unclear which alanine or proline residue in SEQ ID NO: 4 is being referenced, because the C-terminal residue of full-length SEQ ID NO: 4 is cysteine (C297). Consequently, it is unclear whether claim 5 is requiring the cellulase component to be at least 80% identical to the version of SEQ ID NO: 4 with alanine or the changed version with proline. Furthermore, claim 6 requires the cellulase component to have at least 80% sequence identity over the residues 22-228 with the SEQ ID NO: 12. SEQ ID NO: 12 includes an alanine residue at position 228 and at the C-terminal, position 235. Therefore, in claim 6 it is unclear which alanine residue is required to be changed to proline, and it is unclear whether that change broadens the scope of the at least 80% sequence identity requirement.
Claim Rejections - 35 USC § 112(d)
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claim 5 is rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 5 fails to include all the limitations of claim 1 upon which it depends. Claim 1 requires the C-terminal amino acid of the cellulase component to be changed from alanine to proline (A228P) (lines 3-4). Claim 5 requires the cellulase component to have at least 80% sequence identity over residues 18-224 with the SEQ ID NO: 4. Since claim 5 does not require the cellulase component to include residue 228, claim 5 fails to include the “(A228P)” recited in claim 1. Furthermore, the C-terminal amino acid of SEQ ID NO: 4 is cysteine, C297, not alanine or proline. As such, claim 5 does not require the cellulase component to include a C-terminal amino acid that is changed from alanine to proline.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 is/are rejected under 35 U.S.C. 103 as being unpatentable over Juntunen (WO 2020/099719, effective filing date of 11/13/2018 and designates U.S.; hereafter Juntunen2018).
The applied reference has common Inventors with the instant application. Based upon the earlier effectively filed date of the reference, it constitutes prior art under 35 U.S.C. 102(a)(2).
Claim 1 is interpreted as requiring a cellulase component that is an endoglucanasae, wherein the C-terminal amino acid of the cellulase component is changed from alanine to proline (A228P). A228 of a cellulase component, such as instantly disclosed SEQ ID NO: 12, may be changed to P.
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[AltContent: textbox (Juntunen2018 teaches DC121 which includes N210S, N215R, N220S and N225S substitutions. See p. 26. Therefore, DC121 includes a subsequence that is 100% identical to instant SEQ ID NO: 1)]Regarding claim 1, Juntunen2018 teaches variants that are fusion proteins. See p. 12 lines 1-2. Juntunen2018 teaches SEQ ID NO: 1, which is a mature ACM88 cellulase derived from Acremonium thermophilum GH45 cellulase. See p. 5 lines 12-14. GH45 refers to the glycosyl hydrolase family 45 of endoglucanases. See p. 5 lines 21-22. Juntunen2018 teaches that ACM88 contains a Cel45A catalytic core derived from A. thermophilum (i.e. cellulase that is an endoglucanase) attached to a linker and CBM (i.e. cellulose binding). See p. 11 lines 1-3. CBM stands for carbohydrate binding module, which may also be referred to as a cellulose binding domain. See 12 lines 10-12. In example 1, Juntunen2018 teaches producing ACM88 cellulase variants. See p. 11 lines 20-22. These variants include the DC121 variant, which includes N210S, N215R, N220S and N225S substitutions. See table 1 p. 26. An alignment between instant SEQ ID NO: 1 linker (top sequence) and Juntunen2018’s ACM88 SEQ ID NO: 1 (bottom sequence) is shown below.
Juntunen2018’s DC121 variant is 100% identical to instant SEQ ID NO: 1, because the N210S substitution corresponds to a S residue at position 3 of instant SEQ ID NO: 1, the N215R substitution corresponds to an R residue at position 8 of instant SEQ ID NO: 1, the N220S substitution corresponds to a S residue at position 13 of instant SEQ ID NO: 1, and the N225 substitution corresponds to a S residue at position 18 of instant SEQ ID NO: 1. Juntunen2018 suggests adding the variants to liquid used in treating fabric containing cellulose or cellulose derivative. See claim 23 of Juntunen2018. The variants have antipilling and improved stability in the presence of proteases. See the abstract.
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[AltContent: textbox (Instant SEQ ID NO: 12 has an A228 residue, which is changed to proline at the corresponding residue in Juntunen2018’s SEQ ID NO: 1)]Juntunen2018 does not explicitly teach a C-terminal amino acid of the cellulase component that is changed from alanine to proline (A228P); however, as shown in the alignment below, Juntunen2018’s SEQ ID NO: 1 includes the substitution A228P relative to instant SEQ ID NO: 12, a cellulase (specification p. 6, lines 21-23).
Juntunen2018 does not teach a fusion protein that has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 14 or SEQ ID NO: 14, in a liquid detergent in the presence of a protease.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention that Juntunen2018’s variant fusion protein (DC121) necessarily has an at least 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 14 or SEQ ID NO: 14, in a liquid detergent in the presence of a protease. Juntunen2018’s DC121 variant meets all of the instantly claimed structural limitations. MPEP 2112.01(II) states that "products of identical chemical composition cannot have mutually exclusive properties." In re Spada, 911 F.2d 705, 709, 15 USPQ2d 1655, 1658 (Fed. Cir. 1990).
Regarding claim 4, Juntunen2018 teaches that ACM88 contains a Cel45A catalytic core derived from A. thermophilum. See p. 11 lines 1-3. Juntunen2018 teaches fungal endoglucanases belonging to glycosyl hydrolase family 45 (GH45), especially to variants of these endoglucanases. Variants of Acremonium thermophilum Cel45A endoglucanase polypeptide are disclosed. See p. 5 lines 22-26.
Regarding claim 5, Juntunen2018 (p. 5 lines 12-14) teaches SEQ ID NO: 1 including a subsequence that is a 100% identity match to instant SEQ ID NO: 4 residues 18-224. See the alignment below and in the office action appendix.
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[AltContent: textbox (Instant SEQ ID NO: 4 residues 18-224 (top sequence) and Juntunen2018’s SEQ ID NO: 1 (bottom sequence))]
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[AltContent: textbox (Instant SEQ ID NO: 12 residues 22-228 (top sequence) and Juntunen2018’s SEQ ID NO: 1 (bottom sequence))]Regarding claim 6, Juntunen2018 (p. 5 lines 12-14) teaches SEQ ID NO: 1 including a subsequence that is an 80.9% identity match to instant SEQ ID NO: 12 residues 22-228. See the alignment below and in the office action appendix.
Regarding claim 7, Juntunen2018 (p. 5 lines 12-14) teaches SEQ ID NO: 1 including a 38-residue linker from residue 208-245, which is at least 35 amino acids as instantly required. See the alignment between the linker instant SEQ ID NO: 1 and Juntunen2018’s SEQ ID NO: 1 residues 208-245 above.
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[AltContent: textbox (Instant SEQ ID NO: 3 (top sequence) and Juntunen2018’s SEQ ID NO: 1 (bottom sequence))]Regarding claim 9, Juntunen2018 teaches SEQ ID NO: 1. See p. 5 lines 12-14. Juntunen2018 teaches G219S and G244T substitutions. See p. 7 lines 25-26. As shown in the alignment below, residue 219 of Juntunen2018’s SEQ ID NO: 1 corresponds to residue 12 of instant SEQ ID NO:3 linker, and residue 244 of Juntunen2018’s SEQ ID NO: 1 corresponds to residue 37 of instant SEQ ID NO: 3 linker.
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[AltContent: textbox (SEQ ID NO: 14 residues 269-304 (top sequence) and Juntunen’s SEQ ID NO: 1 (bottom sequence))]Regarding claim 11, Juntunen2018 (p. 5 lines 12-14) teaches SEQ ID NO: 1 including a subsequence that is 100% identical to residues 269-304 of instant SEQ ID NO: 14. See the alignment below and the office action appendix.
[AltContent: textbox (Left: Image of each substitution taught or suggested by Juntunen2018;
Right: alignment of modified Juntunen2018’s SEQ ID NO: 1 (top), and instant SEQ ID NO: 16 (bottom))]Regarding claim 12, Juntunen2018 teaches SEQ ID NO: 1. See p. 5 lines 12-14. Juntunen2018’s SEQ ID NO: 1 is an 80.5% identity match with instant SEQ ID NO: 16. Juntunen2018 teaches S23P, A77S, N210S, N215R, N220S, and N225S substitutions. See claim 8 of Juntunen2018. Furthermore, Juntunen2018 suggests substitutions at positions N42, K44, T59, Q82, E116, I130, S136, and T180. See table 1. Juntunen2018 teaches Q58H, D151S, and T193V substitutions. See table 1.
Juntunen2018 does not explicitly teach a fusion protein having at least 90% sequence identity with SEQ ID NO: 16, 18 or 20.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to modify Juntunen2018’s SEQ ID NO: 1 by combining the S23P, A77S, N210S, N215R, N220S, N225S, Q58H, D151S, and T193V substitutions, and by further substitute any amino acid for the N42, K44, T59, Q82, E116, I130, S136, and T180 residues taught by Juntunen2018 based on Juntunen2018’s suggestions. One of ordinary skill in the art in the art would have been motivated to combine the substitutions taught by Juntunen2018, because Juntunen2018 suggests that the variants show improved stability in detergent when compared to the parental enzyme to which substitutions are made. See p. 2 lines 6-8. There would have been a reasonable expectation of success because Juntunen2018 demonstrates varying the parental enzyme to include the S23P, A77S, N210S, N215R, N220S, N225S, Q58H, D151S, and T193V. One of ordinary skill in the art would have been further motivated to substitute any amino acid at positions N42, K44, T59, Q82, E116, I130, S136, and T180, because Juntunen2018 suggests substituting SEQ ID NO: 1 at those positions. There would have been a reasonable expectation of success because Juntunen2018 demonstrates varying SEQ ID NO: 1 at the N42, K44, T59, Q82, E116, I130, S136, and T180; and there are a finite number of amino acids to choose from.
Regarding claim 13, Juntunen2018 teaches an isolated nucleic acid molecule comprising a nucleotide sequence which encodes the variant. See claim 9 of Juntunen2018. Juntunen2018 teaches a variant that comprises N210S, N215R, N22S and N225S substitutions. See claim 8 of Juntunen2018. The DC121 variant, which includes N210S, N215R, N220S and N225S substitutions, includes a subsequence that is 100% identical to instant SEQ ID NO: 1. See table 1 p. 26, and the alignment between instant SEQ ID NO: 1 and Juntunen2018’s SEQ ID NO: 1.
Regarding claim 14, Juntunen2018 teaches a recombinant expression vector comprising the isolated nucleic acid molecule operably linked to regulatory sequences capable of directing expression of a gene encoding said variant cellulase (i.e. fusion protein) in a suitable host. See claim 10 of Juntunen2018.
Regarding claim 17, Juntunen2018 teaches an enzyme composition comprising the variant. See claim 14 of Juntunen2018. Juntunen2018 teaches the enzyme composition characterized in that said composition further comprises: optionally at least one polyol selected from propylene glycol, glycerol, a sugar, sugar alcohol, sorbitol, hexylene glycol; optionally at least one preservative selected from organic acids; optionally at least one inhibitor selected from formic acid, lactic acid, boric acid, boric acid derivative, aromatic borate ester, phenyl boronic acid de rivative, peptide, other reversible subtilisin inhibitors or a combination thereof; optionally at least one enzyme selected from proteases, amylases, cellu- lases, lipases, xylanases, mannanases, cutinases, esterases, phytases, nucleases, pectinases, pectinolytic enzymes, pectate lyases, carbohy- drases, arabinases, galactanases, xanthanases, xyloglucanase, lac- cases, peroxidases and oxidases with or without a mediator, or a combi nation thereof; optionally at least one salt selected from sodium chloride, potassium chloride, potassium (hydrogen)phosphate, sodium (hydrogen)phosphate, ammonium sulfate, potassium sulfate, or a combination thereof; and optionally at least one filler or carrier selected from maltodextrin, flour, sodium chloride, sulfate, sodium sulfate, sodium acid pyrophosphate, tetrasodium pyrophosphate, polyethylene glycol, or a combination there of.
Regarding claim 18, Juntunen2018 teaches an enzyme composition that is in the form of liquid composition or a solid composition. See claim 16 of Juntunen2018.
Regarding claim 19, Juntunen2018 teaches a detergent composition comprising the variant (i.e. fusion protein). See claim 17 of Juntunen2018. The detergent composition preferably comprises one or more of the surfactants, builders, chleators or chleating agents, bleach system or bleach components, polymers, fabric conditioners, foam boosters, sud suppressors, dyes, perfume, tannish inhibitors, optical brighteners, bactericides, fungicides, soil suspending agents, anticorrosion agents, hydrotropes, fabric hueing agents, dispersants, dye transfer inhibiting agents, fluorescent whitening agents, soil release polymers, anti-redepositions agents, anti-shrink agents, anti-wrinkling agents, bactericides, binders, carriers, dyes, enzyme stabilizers, fabric softeners, fillers, foam regulators, perfumes, pigments, buffers, preservatives, sod suppressors, solvents, and structurants for liquid detergents, structure elasticizing agents, enzyme inhibitors or stabilizers, enzyme activators, transferase(s), hydrolytic enzymes, oxido reductases, bluing agents and fluorescent dyes, antioxidants, and solubilizers. See claim 19 of Juntunen2018.
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[AltContent: textbox (Instant SEQ ID NO: 3 (top sequence) and Juntunen2018’s SEQ ID NO: 1 (bottom sequence))]Regarding claim 27, Juntunen2018 teaches SEQ ID NO: 1. See p. 5 lines 12-14. Juntunen2018 teaches G219S and G244T substitutions. See p. 7 lines 25-26. As shown in the alignment below, Juntunen2018’s G219S substitution corresponds to S at position 12 of instant SEQ ID NO: 3 linker and the G244T substitution corresponds to T at position 37 of instant SEQ ID NO: 3 linker.
Regarding claim 28, Juntunen2018 teaches an isolated nucleic acid molecule comprising a nucleotide sequence which encodes the variant (i.e. fusion protein). See claim 9 of Juntunen2018. Juntunen2018 teaches SEQ ID NO: 1, which is the amino acid sequence of mature ACM88. See p. 5 lines 12-13. Juntunen2018 teaches SEQ ID NO: 3, which is the nucleotide sequence of the full-length ACM88 cellulase. See p. 5 lines 18-19. In example 1, Juntunen2018 teaches constructing expression plasmids for the production of recombinant ACM88 variants. See p. 23 lines 1-2. Juntunen2018 teaches the cellulase variants in table 1, and table 1 teaches and suggests the following substitutions: S23P, A77S, N210S, N215R, N220S, N225S, Q58H, D151S, T193V, N42, K44, T59, Q82, E116, I130, S136, and T180.
Juntunen2018 does not teach an isolated nucleic acid encoding the fusion protein having at least 90% sequence identity with SEQ ID NO: 16, 18 or 20.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to modify Juntunen2018’s SEQ ID NO: 3 as to arrive at a nucleotide sequence encoding the variant (discussed above with respect to instant claim 12) with the following substitutions taught or suggested by Juntunen2018: S23P, A77S, N210S, N215R, N220S, N225S, Q58H, D151S, T193V, N42, K44, T59, Q82, E116, I130, S136, and T180. One of ordinary skill in the art in the art would have been motivated to do so, because Juntunen2018 suggests that such variants show improved stability in detergent when compared to the parental enzyme to which substitutions are made. See p. 2 lines 6-8. There would have been a reasonable expectation of success because Juntunen2018 demonstrates constructing expression plasmids (i.e. nucleic acid sequences) of recombinant ACM88 in order to produce variants.
Regarding claim 29, Juntunen2018 teaches a recombinant expression vector comprising the isolated nucleic acid molecule operably linked to regulatory sequences capable of directing expression of a gene encoding said variant cellulase (i.e. fusion protein) in a suitable host. See claim 10 of Juntunen2018.
Regarding claim 37, Juntunen2018 teaches a detergent composition in the form of a liquid detergent or a solid detergent. See claim 18 of Juntunen2018. Juntunen2018 teaches a detergent composition comprising one or more additional enzymes selected from the group of proteases, amylases, cellulases, lipases, xylanases, mannanases, cutinases, esterases, nucleases, pectinases, pectate lyases, pectinolytic enzymes, carbohydrases, arabinases, galactanases, xanthanases, xyloglucanases, laccases, peroxidases and oxidases, and wherein the detergent composition preferably comprises one or more of the surfactants, builders, chelators or chelating agents, bleach system or bleach components, polymers, fabric conditioners, foam boosters, suds suppressors, dyes, perfume, tannish inhibitors, optical brighteners, bactericides, fungicides, soil suspending agents, anticorrosion agents, hydrotropes, fabric hueing agents, dispersants, dye transfer inhibiting agents, fluorescent whitening agents, soil release polymers, anti-redepositions agents, anti-shrink agents, anti-wrinkling agents, bactericides, binders, carriers, dyes, enzyme stabilizers, fabric softeners, fillers, foam regulators, perfumes, pigments, buffers, preservatives, sod suppressors, solvents, and structurants for liquid detergents, structure elasticizing agents, enzyme inhibitors or stabilizers, enzyme activators, transferase(s), hydrolytic enzymes, oxido reductases, bluing agents and fluorescent dyes, antioxidants, and solubilizers. See claim 19 of Juntunen2018.
Regarding claim 38, J Juntunen2018 teaches a detergent composition, characterized in that it comprises the variant or the enzyme composition. See claim 38 of Juntunen2018.
Regarding claim 39, Juntunen2018 teaches a detergent composition in the form of a liquid detergent or a solid detergent. See claim 18 of Juntunen. Juntunen teaches a detergent composition comprising one or more additional enzymes selected from the group of proteases, amylases, cellulases, lipases, xylanases, mannanases, cutinases, esterases, nucleases, pectinases, pectate lyases, pectinolytic enzymes, carbohydrases, arabinases, galactanases, xanthanases, xyloglucanases, laccases, peroxidases and oxidases, and wherein the detergent composition preferably comprises one or more of the surfactants, builders, chelators or chelating agents, bleach system or bleach components, polymers, fabric conditioners, foam boosters, suds suppressors, dyes, perfume, tannish inhibitors, optical brighteners, bactericides, fungicides, soil suspending agents, anticorrosion agents, hydrotropes, fabric hueing agents, dispersants, dye transfer inhibiting agents, fluorescent whitening agents, soil release polymers, anti-redepositions agents, anti-shrink agents, anti-wrinkling agents, bactericides, binders, carriers, dyes, enzyme stabilizers, fabric softeners, fillers, foam regulators, perfumes, pigments, buffers, preservatives, sod suppressors, solvents, and structurants for liquid detergents, structure elasticizing agents, enzyme inhibitors or stabilizers, enzyme activators, transferase(s), hydrolytic enzymes, oxido reductases, bluing agents and fluorescent dyes, antioxidants, and solubilizers. See claim 19 of Juntunen.
Regarding claim 43, Juntunen2018 teaches an enzyme composition comprising at least one preservative selected from organic acids, e.g. benzoic acid, citric acid, ascorbic acid, sorbic acid, and salts thereof, sodium benzoate, hydroxybenzoate, benzisothiazolinone (BIT) or a combination thereof. See claim 15 of Juntunen.
Regarding claim 44, Juntunen2018 teaches an enzyme composition characterized in that said enzyme composition is in the form of liquid composition or a solid composition such as solution, dispersion, paste, powder, pellet, granule, granulate, coated granulate, tablet, cake, crystal, crystal slurry or gel. See claim 16 of Juntunen.
This rejection under 35 U.S.C. 103 might be overcome by: (1) a showing under 37 CFR 1.130(a) that the subject matter disclosed in the reference was obtained directly or indirectly from the inventor or a joint inventor of this application and is thus not prior art in accordance with 35 U.S.C.102(b)(2)(A); (2) a showing under 37 CFR 1.130(b) of a prior public disclosure under 35 U.S.C. 102(b)(2)(B); or (3) a statement pursuant to 35 U.S.C. 102(b)(2)(C) establishing that, not later than the effective filing date of the claimed invention, the subject matter disclosed and the claimed invention were either owned by the same person or subject to an obligation of assignment to the same person or subject to a joint research agreement. See generally MPEP § 717.02.
Claims 1, 3-7, 9, 11, 13-14, 17-19, 27, 37-39, and 44 are rejected under 35 U.S.C. 103 as being unpatentable over Juntunen (WO 2016/066896), in view of Schnorr (US 9,765,373, published 09/19/2017), Payne (Proceedings of the National Academy of Sciences, 2013110(36), 14646-14651), and Montalibet (US 2013/0052694).
Regarding claim 1, Juntunen teaches a cel45A-ACM0 cellulase variant, corresponding to SEQ ID NO: 12, which contains a catalytic core of A. thermophilum Cel45A (i.e. cellulase that is an endoglucanase) attached to a linker and a carbohydrate binding module (CBM) region of T. reesei Cel7A. See example 1, specifically the paragraph spanning pages 16-17, and claims 1-3. Juntunen discloses that cellulases are used to prevent formation of pills (e.g. antipilling) on the surface of cotton garments (e.g. fabric containing cellulose). See lines 24-28 on page 1 and lines 28-34 on page 14. Juntunen discloses that carbohydrate binding modules or domains (CBM/CBD) enhance the binding of cellulase enzymes to cellulose-containing fiber. See page 1 lines 15-23 and page 8 lines 31-36. Moreover, Juntunen teaches a linker connecting the CBM and catalytic domain via a flexible and highly glycosylated region. See lines 3-4 of page 9. Residues 225-262 of SEQ ID NO: 12 are a 93.3% identity match to the instant SEQ ID NO: 1 linker. Therefore, Juntunen teaches a cel45A-ACM0 (SEQ ID NO: 12) fusion protein containing an endoglucanase cellulase component (i.e. the A. thermophilum Cel45A), a linker component (i.e. residues 225-262 of SEQ ID NO: 12) and a T. reesei Cel7A carbohydrate binding module region that can bind to cellulose; and wherein the cellulase has an antipilling effect on fabric containing cellulose.
Juntunen does not teach a cellulose component, wherein the C-terminal amino acid of the cellulase component is changed from alanine to proline (A228).
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The instant specification discloses that the core region of a cellulase component according to the invention corresponds to the amino acids 18-224 of SEQ ID NO: 4. See p. 8 lines 10-13.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention that the C-terminal residue of the cellulase component in Juntunen’s SEQ ID NO: 12 is a proline.
Juntunen does not teach a linker component that has at least a 94% sequence identity with SEQ ID NOs: 1, 2, or 3, and, according to the numbering of SEQ ID NO: 1, comprises an amino acid at position 3 that is S; an amino acid at position 8 that is R; an amino acid at position 13 that is S; and an amino acid position 18 that is S.
Schnorr teaches glycoside hydrolase polypeptide comprising a carbohydrate binding module and a linker. See lines 21-50 in column 60. Schnorr teaches examples of common substitutions including Ser/Asn. See column 16 lines 5-10.
Payne teaches that there is little sequence conservation in linkers that connect carbohydrate binding modules and catalytic domains in lignocellulose-degrading enzymes. The linkers typically contain significant glycine, proline, serine and threonine content. The serine and threonine residues often exhibit O-glycosylation, which imparts protease resistance. See the first two paragraphs on page 14646. Payne discloses that linker glycosylation is known to provide protease protection, and that linker domains in many lignocellulose-degrading enzymes are likely optimized in length and sequence to play a more direct role in interacting with plant cell walls. See the conclusion section on page 14650.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to replace all of the asparagine (N) residues in the linker region spanning residues 225-262 of Juntunen’s SEQ ID NO: 12 with serine (S), in view of Schnorr and Payne. One of ordinary skill in the art would have been motivated to do so because Juntunen suggests using a highly glycosylated linker region, and Payne suggests that serine residues often exhibit O-glycosylation, which imparts protease resistance (paragraph 2 page 14647). There would be a reasonable expectation of success because Schnorr suggests that asparagine is interchangeable with serine in proteins having cellulolytic activity.
Juntunen, Schnorr and Payne do not teach a linker component that has at least a 94% sequence identity with SEQ ID NOs: 1, 2, or 3, and, according to the numbering of SEQ ID NO: 1, comprises an an amino acid at position 8 that is R.
Montalibet teaches an isolated Trichoderma reesei TrCel7A cellobiohydrolase comprising one or more amino acid substitutions selected from a list that includes N431X, wherein the isolated Trichoderma reesei TrCel7A cellobiohydrolase comprises an amino acid sequence that is from about 75% to about 99.9% identical to amino acids 1-497 of SEQ ID NO: 1 and exhibits: (a) increased specific activity, (b) reduced inhibition by glucose, (c) reduced inactivation by lignin, (d) increased activity in the presence of lignin, (e) increased activity in the presence of lignocellulose hydrolysate, or (f) an combination of (a) through (e), relative to a parental Trichoderma reesei TrCel7A. See claim 8. Montalibet teaches the isolated Trichoderma reesei TrCel7A cellobiohydrolase of claim 8 comprising one or more amino acid substitution selected from a group that includes N431R. See claims 9 and 10. Montalibet teaches SEQ ID NO: 146, which is the isolated TrCel7A associated with the N431R substitution. See table 3. The N431R substitution of Montalibet aligns with residue 8 of instant SEQ ID
NO: 1. See the office action appendix for the full multiple sequence alignment.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to replace the asparagine (N) at position 232 of Juntunen’s SEQ ID NO: 12 with an arginine (R) residue based on Montalibet’s suggestion. One of ordinary skill in the art would have been motivated to do so because Montalibet suggests that the N431R mutation is associated with a) increased specific activity, (b) reduced inhibition by glucose, (c) reduced inactivation by lignin, (d) increased activity in the presence of lignin, and/or (e) increased activity in the presence of lignocellulose hydrolysate. There would have been a reasonable expectation of success because the N431R substitution is the only difference between residues 430-497 of Montalibet’s SEQ ID NO: 146 and residues 231-298 of Juntunen’s SEQ ID NO: 12. As shown in the alignment below, the modified of Juntunen, Schnorr, Payne, and Montalibet is 100% identical to instant SEQ ID NO: 1 and includes, relative to instant SEQ ID NO: 1, a S at positions 3, a R at position 8, a S at position 13 and an S at position 18.
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[AltContent: textbox (Alignment between from top to bottom, SEQ ID NO: 146 of “Motalibet” [sic Montalibet], Juntunen SEQ ID NO: 12 and instant SEQ ID NO: 1. The underlined portion of Juntunen SEQ ID NO: 12 corresponds with the linker region (residues 225-262). The boxes show the residue substitutions discussed in the rationale. From left to right the boxes show: the N227 residue of Juntunen’s SEQ ID NO: 12, residue 431 in Montalibet’s SEQ ID NO: 146, and residues N237 and N242 of Juntunen’s SEQ ID NO: 12. )]Juntunen, Schnorr, Payne and Montalibet do not teach a fusion protein that has at least 5-fold increase in improved protease stability compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14 in a liquid detergent in the presence of a protease.
Based on the instant claim language, the instantly claimed improved protease stability is inherent to the instantly claimed fusion protein structure.
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to recognize that the modified fusion protein of Juntunen, Schnorr, Payne and Montalibet is capable of having an at least 5-fold increase in improved protease stability compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14 in a liquid detergent in the presence of a protease. Juntunen, Schnorr, Payne and Montalibet meet every claimed structural limitation. MPEP 2112.01(II) states that "products of identical chemical composition cannot have mutually exclusive properties." In re Spada, 911 F.2d 705, 709, 15 USPQ2d 1655, 1658 (Fed. Cir. 1990).
Regarding claim 4, Juntunen teaches that the variants of the Acremonium thermonophilum Cel45A endoglucanase peptides belong to the glycosyl hydrolase family 45. See the first full paragraph on page 6.
Regarding claim 5, SEQ ID NO: 12 of Juntunen is a 96.7% identity match over residues 18-224 of instant SEQ ID NO: 4.
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For clarity, the modified SEQ ID NO: 12 of Juntunen, Schnorr, Payne, and Montalibet as discussed above regarding claim 1, is also 96.7% identical to residues 18-224 of instant SEQ ID NO: 4. See the alignment below between residues 18-224 of instant SEQ ID NO: 4 (top) and residues 18-224 of SEQ ID NO: 12 of Juntunen, as modified by Schnorr, Payne and Montalibet (bottom).
Regarding claim 6, SEQ ID NO: 12 of Juntunen is an 80.9% identity match over residues 22-228 of instant SEQ ID NO: 12.
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For clarity, residues 19-223 of modified SEQ ID NO: 12 of Juntunen, Schnorr and Payne, as discussed above regarding claim 1, is also 80.9% identical to residues 22-228 of instant SEQ ID NO: 12.
Regarding claim 7, residues 225-262 of SEQ ID NO: 12 of Juntunen are a 93.3% identity match to instant SEQ ID NO: 1. Therefore, Juntunen teaches a linker that is 37 residues in length (i.e. 262-225=37), which is at least 35 amino acids in length, as instantly claimed.
Regarding claims 9 and 27, Juntunen teaches variant polypeptides having an amino acid substitution, deletion or insertion. See page 8 paragraph 1. Juntunen teaches SEQ ID NO: 12, wherein residues 225-262 of are a 93.3% identity match to instant SEQ ID NO: 1.
Juntunen does not teach an amino acid sequence of a linker component further comprising at least the following characteristics: the amino acid at position 12 is not G, and the amino acid position at position 37 is not G, with respect to the position number of instant SEQ ID NO: 1 (relevant to instant claim 9), wherein the amino acid position at position 12 is S and the amino acid position at position 37 is T (relevant to instant claim 27).
Schnorr suggests making amino acid substitutions, such as substitutions amongst small amino acids including glycine, serine and threonine. See column 16 line 3-4.
Payne teaches that there is little sequence conservation in linkers that connect carbohydrate binding modules and catalytic domains in lignocellulose-degrading enzymes. The linkers typically contain significant glycine, proline, serine and threonine content. The serine and threonine residues often exhibit O-glycosylation, which imparts protease resistance. See the first two paragraphs on page 14646.
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Montalibet discloses that linker peptides are typically basic peptides, particularly enriched in serine, threonine and proline. Montalibet indicates that proline, serine and threonine may account for 50% or more of the amino acids in all linker peptide sequences from bacterial and fungal glycoside hydrolases, e.g. endoglucanases. See paragraph [0061].
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to further replace any of the glycine(G) residues in the modified linker region of Juntunen discussed above with serine(S) or threonine(T) in view of Schnorr, Payne and Montalibet. Doing so is mere substitution, and pursuing the known options within one’s technical grasp. In the process, one could reasonably arrive at a linker region comprising a G12T or G12S substitution, and a G37T substitution relative to instant SEQ ID NO: 1. See the marked residues in the image above. One would be motivated to use serine and threonine residues in the linker region because serine and threonine may contribute to protease resistance, according to Payne (paragraph 2 page 14647). There would be a reasonable expectation of success because Schnorr suggests that glycine is interchangeable with serine or threonine in proteins having cellulolytic activity. Moreover, a person of ordinary skill in the art has good reason to pursue the known options within his or her technical grasp. One could reasonably substitute the glycine residues at positions 236 and 261 of SEQ ID NO: 12 because there are only 8 glycine residues within the linker region of Juntunen’s SEQ ID NO: 12 to choose from.
Regarding claim 11, residues 263-298 in SEQ ID NO: 12 of Juntunen are a 100% identity match over residues 269-304 of instant SEQ ID NO: 14. See the alignment below. Note that residues 263-298 in SEQ ID NO: 12 of Juntunen are not within the linker region.
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Regarding claim 13, Juntunen teaches cel45A-ACM0, corresponding to nucleic acid sequence SEQ ID NO: 3 and amino acid sequence SEQ ID NO: 12. The cel45A-ACM0 variant, contains a catalytic core of A. thermophilum Cel45A attached to a linker and a carbohydrate binding module (CBM) region of T. reesei Cel7A. See the paragraph spanning pages 16-17 and claims 1-3.
Regarding claim 14, Juntunen teaches constructing expression plasmids. The constructs contain a T. reesei cel7A promoter and a terminator. See the paragraph spanning pages 16-17. In claims 10-12, Juntunen teaches a variant endoglucanase polypeptide that has an amino acid sequence SEQ ID NO: 12; a polynucleotide encoding the polypeptide and a vector which comprises the polynucleotide operably linked to regulatory sequences capable of directing expression of the polypeptide.
Regarding claim 17, Juntunen teaches a cel45A-ACM0 (SEQ ID NO: 12) containing a linker and CBM. See the paragraph spanning pages 16-17 and claims 1-3. Juntunen teaches using suitable stabilizers including polyols, such as propylene, glycol, glycerol, a sugar or sugar alcohol. See page 12 lines 28-29. Juntunen teaches using preservatives. See claim 17 of Juntunen. Juntunen teaches using lactic acid, boric acid, or boric acid derivatives (e.g. inhibitors). See page 12 lines 28-29. Juntunen teaches using enzymes such as proteases, amylases, other cellulases, lipases, pectinolytic enzymes. See claim 16 of Juntunen. Detergent compositions may also comprise poly(ethylene glycol) (e.g. a filler or carrier). See line 3 of page 13.
Regarding claims 18 and 44, Juntunen teaches that the enzyme preparations may be provided as a liquid or as a solid, for example as a dried powder, a tablet, a crystal or crystal slurry, See the first full paragraph on page 12. The endoglucanase polypeptide variant or an enzyme preparation thereof can be in the form of a gel or in granules. See page 13 lines 28-29.
Regarding claims 19, Juntunen teaches an endoglucanase polypeptide variant or an enzyme preparation thereof with surfactants, builders, bleaching agents (e.g. bleach component), polymers/antiredeposition agents (e.g. polymers and anti-redepositions agents), dyes, perfumes, optical brighteners, and anti-corrosion agents (page 3 lines 23-28).
Regarding claim 37, Juntunen teaches enzyme preparations (e.g. a detergent) that may be provided as a liquid or solid. See the first full paragraph on page 12. Enzyme preparations may further comprise at least one enzyme selected from a group that includes proteases, amylases, other cellulases, lipases, and pectinolytic enzymes. See claim 16 of Juntunen. Furthermore, Juntunen teaches enzyme preparations with surfactants, builders, bleaching agents (e.g. bleach component), polymers/antiredeposition agents (e.g. polymers and anti-redepositions agents), dyes, perfumes, optical brighteners, and anti-corrosion agents (page 3 lines 23-28).
Regarding claim 38, Juntunen teaches detergent compositions comprising endoglucanase polypeptide variants. See claim 17 of Juntunen.
Regarding claim 39, Juntunen teaches detergent compositions (e.g. see claim 17 of Juntunen) and using enzymes such as proteases, amylases, other cellulases, lipases, pectinolytic enzymes (claim 16 of Juntunen). Juntunen teaches enzyme preparations with surfactants, builders, bleaching agents, polymers/antiredeposition agents, dyes, perfumes, optical brighteners, and anti-corrosion agents (page 3 lines 23-28).
Claims 12, 28-29 and 43 are rejected under 35 U.S.C. 103 as being unpatentable over Juntunen (WO 2016/066896), Schnorr (US 9,765,373, published 09/19/2017), Payne (Proceedings of the National Academy of Sciences, 2013110(36), 14646-14651), and Montalibet (US 2013/0052694), as applied to claims 1, 3, 5-7, 9, 11, 13-14, 17-19, 26-27, 37-39, and 44 above, and further in view of Chieffi (EP 3301148 A1) with evidence from Arola, (2016) Scientific Reports, 6(1), 1-9.
Juntunen, Schnorr, Payne and Montalibet render obvious instant claim 1, as discussed above.
Regarding claim 12, Juntunen teaches a linker connecting the CBM and catalytic domain via a flexible and highly glycosylated region. See lines 3-4 of page 9. Residues 225-262 of SEQ ID NO: 12 are a 93.3% identity match to the instant SEQ ID NO: 1 linker.
Schnorr teaches common substitutions, such as Ser/Asn. See column 16 lines 8-10.
Payne suggests that serine residues often exhibit O-glycosylation, which imparts protease resistance. See the first two paragraphs on page 14646.
Montalibet teaches an isolated Trichoderma reesei TrCel7A cellobiohydrolase of comprising one or more amino acid substitution selected from a group that includes N431R. See claims 9 and 10. Montalibet teaches SEQ ID NO: 146, which is the isolated TrCel7A associated with the N431R substitution. See table 3. The N431R substitution of Montalibet aligns with residue 8 of instant SEQ ID NO: 1.
Juntunen, Schnorr, Payne and Montalibet do not teach a fusion protein that is at least 90% identical to instant SEQ ID NOs: 16, 18 or 20.
Chieffi teaches a cellulose that is a hybrid fusion endoglucanase comprising a glycosyl hydrolase family 45 (GH 45) catalytic domain that is a wild-type or variant of a microbially-derived endoglucanase endogenous to Melanocarpus albomyces, and a carbohydrate binding molecule that is a wild type or variant of a carbohydrate binding module endogenous to Trichoderma reesei, and which has at least a 90% identity to amino acid sequence SEQ ID NO: 4. See claim 19 part (p) of Chieffi. The carbohydrate binding molecule of Chieffi is inherently capable of binding to cellulose because evidentiary reference Arola discloses that T. reesei CBMs bind cellulose. See the abstract, figure 1, and the last paragraph on page 2 of Arola. SEQ ID NO: 4 of Chieffi is a 90.7% identity match to instant SEQ ID NO: 16, a 90.3% identity match to instant SEQ ID NO: 18, and a 90% identity match to instant SEQ ID NO: 20. Furthermore, Chieffi’s SEQ ID NO: 4 includes a P residue corresponding to residue 224 of instant SEQ ID NO: 4, which is a cellulase domain. Therefore, Chieffi’s SEQ ID NO: 4 meets the instant requirement for the cellulase component to have the C-terminal residue changed from alanine to proline. See the office action appendix for the alignment.
Chieffi meets the limitation of instant claim 12 requiring the fusion protein to have at least a 90% identity match with SEQ ID NO: 16, 18 or 20; but Chieffi does not teach the instantly claimed linker component that is at least 94% identical to instant SEQ ID NO: 1, 2 or 3, and, according to the numbering of instant SEQ ID NO: 1, comprises an amino acid at position 3 that is not N; an amino acid at position 8 that is R; an amino acid at position 13 that is not N; and an amino acid position 18 that is not N (relevant to instant claim 1, from which claim 12 depends).
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to replace the linker within SEQ ID No: 4 of Chieffi with the modified linker of Juntunen, Schnorr, Payne and Montalibet discussed above. Doing so is mere substitution. In the process, one would arrive at a sequence that is 92.5% identical to instant SEQ ID NO: 16, 92.1% identical to instant SEQ ID NO: 18, and 92.1% identical to instant SEQ ID NO: 20. See the alignments provided in the office action appendix. One would be motivated to use the sequence of Chieffi because Chieffi teaches laundry detergent compositions having a low pH profile, good solubility profile, good cleaning profile, good stability profile and good fabric care profile. See paragraph [0001] of Chieffi. One would be motivated to use the linker region of Juntunen, Schnorr, Payne and Montalibet with Cheiffi’s SEQ ID NO: 4 because Juntunen suggests that the linker is flexible, highly glycosylated and suitable for linking CBM and catalytic domains. There would be a reasonable expectation of success because Chieffi discloses SEQ ID NO: 4 has an endoglucanase domain and carbohydrate binding module(paragraph [0071]), and Juntunen teaches a linker connecting the CBM and catalytic domain via a flexible and highly glycosylated region (Juntunen lines 3-4 of page 9).
Regarding claim 28, Juntunen teaches an isolated polynucleotide or complementary DNA encoding an endoglucanase polypeptide variant (e.g. SEQ ID NO: 12). See claim 11 of Juntunen. Juntunen teaches producing cellulase variants from cel45A-ACM0, corresponding to nucleic acid sequence SEQ ID NO: 3 and amino acid sequence SEQ ID NO: 12. See the paragraph spanning pages 16-17 and claims 1-3.
Regarding claim 29, Juntunen teaches constructing expression plasmids. The constructs contain a T. reesei cel7A promoter and a terminator (e.g. regulatory sequences). See the paragraph spanning pages 16-17. In claims 10-12, Juntunen teaches a polynucleotide encoding a polypeptide and a vector which comprises the polynucleotide operably linked to regulatory sequences capable of directing expression of the polypeptide.
The teachings of Juntunen with respect to instant claim 17, from which claim 43 depends, are discussed above.
Regarding claim 43, Juntunen teaches detergent compositions with preservatives for preventing spoilage of other ingredients. See page 13 lines 2-5. Juntunen teaches builders or complexing agents such as citrate (e.g. an anion in salts from citric acid). See line 36 on page 12 to line 2 on page 13.
Chieffi teaches detergent ingredients such as the co-builders citric acid and citrate. See paragraph [0089].
Response to Arguments
Applicant's arguments filed 04/24/2026 have been fully considered, but they are unpersuasive.
Rejection of claims 1, 3-7, 9, 11, 13-14, 17-19, 27, 37-39 and 44 under § 103 over Juntunen, Schnorr, Payne, and Montalibet
Applicant argues that in the Advisory Action, Examiner considers that the claimed fusion protein is allegedly identical with the structure taught by Juntunen, Schnorr, Payne and Montalibet, and that “if desired, the enzymatic activity may be concentrated and/or stabilized for storage”. Applicant asserts that this is a misunderstanding of the claims, and the purpose of the invention. See the remarks p. 10 last full paragraph.
This argument is not persuasive, because the advisory action mailed 09/26/2025 is referencing p. 11 lines 10-14 of Juntunen (WO 2016/066896), which states: If desired, the enzyme preparations may be dried, spray-dried or lyophilized, granulated or the enzymatic activity may be otherwise concentrated and/or stabilized for storage. See the advisory action p. 2 paragraph 5. This portion of Juntunen is referenced in response to an argument in the advisory action mailed 09/26/2025 because the portion of Juntunen shows that Juntunen is in the same field of endeavor compared to the instant invention.
Applicant argues that Payne and the other cited documents do not suggest adding a Pro residue in position Ala228, as is done in the present invention. See the remarks p. 11 paragraph three. Applicant argues that there is no “roadmap” in the cited art that would lead a person of ordinary skill in the art to target this specific junction residue for mutation to proline to improve the stability of a fusion protein. See the remarks p. 11 last full paragraph.
This argument is not persuasive because the amendment requiring the cellulase component to have a C-terminal amino acid change from alanine to proline is addressed above. The instant specification indicates that the core region of a cellulase component according to the invention corresponds to the amino acids 18-224 of SEQ ID NO: 4. See p. 8 lines 10-13. Since the P224 residue of SEQ ID NO: 4 corresponds to the P224 residue of Juntunen’s SEQ ID NO: 12, Juntunen teaches the instantly claimed C-terminal amino acid change. Applicant argues that there is no road map that would lead a person of ordinary skill in the art to target this specific junction residue for mutation to proline. However, this argument is unpersuasive because mere recognition of latent properties in the prior art does not render nonobvious an otherwise known invention. In re Wiseman, 596 F.2d 1019, 201 USPQ 658 (CCPA 1979).
Applicant argues that even assuming, arguendo, that Payne generally suggests that glycosylation provides some protease resistance, Payne fails to teach or suggest that the specific combination of the modifications as claimed would yield at least a 5-fold increase in improved protease stability. As shown in Applicant’s specification, the claimed fusion protein was at least 5 times more stable than the reference fusion protein. This high-magnitude, quantitative improvement is not a predicted or inherent result of routine optimization. See the remarks p. 12 paragraph 1.
This argument is not persuasive because it is not commensurate in scope with the instant claims. In example 3, the instant specification teaches that the stability of MA79+CBML measured as application performance is about 5 times better than with MA79+CBM. See p. 32 lines 7-9. The disclosed “about 5 times better” is not equivalent in scope compared to the instantly claimed “at least a 5-fold increase in improved stability” recited in line 13 of claim 1. Furthermore, the instant claims do not limit the fusion protein to the MA79+CBML embodiment disclosed in example 3. MA79+CBML is SEQ ID NO: 18 (p. 7 line 6) and the MA79+CBM is SEQ ID NO: 14 (p. 6 line 26). As such, the results referenced by Applicant are not commensurate in scope with the instant claims.
Applicant argues that to achieve the claimed fusion protein by combining the various disclosures provided in Juntunen, Schnorr, Payne and Montalibet is not possible without using hindsight. See the remarks p. 12 paragraph 2.
In response to Applicant's argument that the examiner's conclusion of obviousness is based upon improper hindsight reasoning, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971). In the instant case, every limitation is accounted for in the prior art, so nothing is gleaned from the instant disclosure.
Applicant argues that a “latent property” of S431R substitution in Montalibet is not something the skilled person could utilize to select this substitution for solving a technical problem of Juntunen polypeptide, or when trying to develop a fusion protein having antipilling/depilling activity and increased stability against protease. The skilled person can select a particular substitution to solve a technical problem only if in the prior art there is an objective reason, such as persuasive experimental evidence or explicit teaching that the modification solves the technical problem, and which thereby guides the skilled person to select this particular substitution. See the remarks p. 12 last full paragraph. In the presence case, the teaching of Montalibet concerning substitution at S431R is insufficient for the skilled person to consider that the technical problem could be solved by the substitution. Montalibet does not teach that this substitution, when applied to Juntunen sequence, would improve antipilling/depilling activity or protease stability. Montalibet is silent about antipilling, depilling or stability against proteases. See the remarks p. 12 the paragraph spanning pages 12-13.
This argument is not persuasive because Montalibet explicitly teaches an N431R substitution (see claim 9 of Montalibet); and Montalibet teaches that an N431X substitution exhibits increased specific activity, reduced inhibition by glucose, reduced inactivation by lignin, increased activity in the presence of lignin, and/or increased activity in the presence of lignocellulose hydrolysate (claim 8 of Montalibet). It is not necessary that the prior art suggest the combination to achieve the same advantage or result discovered by applicant. See, e.g., In re Kahn, 441 F.3d 977, 987, 78 USPQ2d 1329, 1336 (Fed. Cir. 2006).
Applicant argues that Montalibet’s teaching guides the skilled person away from the solution of claim 1: Montalibet discloses in table 9 (column 47) results from a total 51 variants of TrCel7A. The variant SEQ ID NO: 446 having the substitution S431R is at position 49 out of 51 when ranked by their enzyme activity. See the remarks p. 13 first full paragraph. Because Montalibet teaches that S431R is one of the worst performing substitutions, this is not the substitution choice that the skilled person would apply when trying to improve antipilling/depilling effect or protease stability. Applicant argues that Montalibet fails to identify any modification which would improve antipilling/depilling or protease stability. See the remarks p. 13 last full paragraph. The unsuitability of the substitution S431R for enzyme activity performance on cellulase substrate is further evidenced in Montalibet by the fact that this variant is discarded. See the remarks the paragraph spanning p. 13-14.
This argument is not persuasive because the rejection does not cite table 9 of Montalibet at all, and the rejection does not rely on Montalibet’s S431R substitution. Furthermore, one cannot show non-obviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Montalibet is not relied upon for teaching antipilling, depilling or protease stability.
Applicant argues that structural difference between Montalibet and the presently claimed fusion protein is that the amino acid in the substitute position is Ser in Montalibet, whereas in the claimed fusion protein this is N. Montalibet fails to teach that a N>R substitution would have the same, or actually any technical effect as S>R substitution which is the substitution that is made in claim 1. Consequently, the skilled person would have no incentive or expectation of success in making the modification S431R in the Juntunen structure in order to improve antipilling/depilling or stability against proteases. See the remarks p. 13 paragraph 3.
This argument is not persuasive. Claim 1 is a product, so the claim that does not require any active substitution step. Claim 1 requires R to be present at position 8 with respect to instant SEQ ID NO: 1. Juntunen’s SEQ ID NO: 12 has a N residue corresponding to position 8 of instant SEQ ID NO: 1 (i.e. the N232 in Juntunen’s SEQ ID NO: 12). That same N residue (N232) of Juntunen corresponds with the N431R substitution taught by Montalibet. Montalibet further suggests that the N431 substitution can exhibit, for example, increased specific activity (see claim 8 of Montalibet).
Applicant references Juntunen’s alleged teaching that the enzyme activity can “concentrate and/or stabilize for storage”. Applicant asserts that this is a misunderstanding of the teachings of Juntunen and the present invention. The fusion protein of claim 1 is a single polypeptide molecule and the skilled person knows that it is not possible to concentration a single polypeptide molecule to achieve a desired enzyme activity or stability. A composition comprising such a polypeptide may naturally be provided at different concentrations and with select additives but such means are external to the structure of the fusion protein. See the remarks p. 14 first full paragraph.
This argument is not persuasive because the rejection does not rely on any such “concentrate and/or stabilize for storage” teaching of Juntunen, nor does the rejection suggest concentrating a single polypeptide. To the extent that Applicant is referencing the advisory action mailed 09/26/2025, it is unpersuasive. In the advisory action, Juntunen p. 11 lines 10-14 is referenced, because the cited portion suggests that in an enzyme preparation that consists of desired enzymes, the enzymatic activity may be concentrated and/or stabilized for storage. See p. 2 paragraph 5 of the advisory action. This portion of Juntunen is in no way interpreted as concentrating a single polypeptide.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The actual filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/apply/applying-online/eterminal-disclaimer.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-11 and 26 of copending Application No. 17/291,424 (hereafter Juntunen ‘424) in view of Juntunen WO 2016/066896 (referred to as Juntunen), and Chieffi EP 3301148 A1.
Claim 1 of Juntunen ‘424 recites a variant of a GH45 cellulase polypeptide, wherein the variant comprises substitutions in amino acid positions 167, 210, 215, 220 and 225, wherein the amino acid positions are numbered with reference to a parent GH45 cellulase having an amino acid sequence set forth in SEQ ID NO: 1; wherein the variant has at least 80% sequence identity with the parent GH45 Cellulase of SEQ ID NO: 1; and wherein the variant has improved protease stability in comparison to the parent GH45 cellulase.
Claim 4 of Juntunen ‘424 recites the variant of claim 1, wherein substitutions comprise the substitutions Q167Y, N210S, N215R/S, N220S and N225S.
Claim 6 of Juntunen ‘424 recites the variant of claim 1, wherein the variant further comprises one or more further substitutions in the positions 23, 44, 65, 66, 77, 193, 219, 242 and 244.
Claim 7 of Juntunen ‘424 recites the variant of claim 6, wherein the further substitution is one or more of the substitutions S23P, K44N, D65E, N66Q, A77S, T193V, G219S, S242G and G244T.
Claim 8 of Juntunen ‘424 recites the variant of claim 6, comprising the following substitutions: S23P, D65E, N66Q, A77S, Q167Y, N210S, N215R, G219S, N220S, N225S, and G244T.
Claim 9 of Juntunen ‘424 recites an isolated nucleic acid molecule comprising a nucleotide sequence which encodes the variant of claim 1.
Claim 10 of Juntunen ‘424 recites a recombinant expression vector comprising a nucleotide sequence of claim 9 operably linked to regulatory sequences capable of directing expression of the gene encoding said variant cellulase in a suitable host.
Claim 11 of Juntunen ‘424 recites a host cell comprising the recombinant expression vector according to claim 10, the host cell preferably being selected from the group consisting of: fungal cells, filamentous fungal cells from Division Ascomycota, Subdivision Pezizomycotina; Class Sordariomycetes, Subclass Hypocreomycetidae, Orders Hypocreales and Microascales and Aspergillus, Chrysosporium, Myceliophthora and Humicola; Families Hypocreacea, Nectriaceae, Clavicipitaceae, Microascaceae, and Genera Trichoderma (anamorph of Hypocrea), Fusarium, Gibberella, Nectria, Stachybotrys, Claviceps, Metarhizium, Villosiclava, Ophiocordyceps, Cephalosporium, and Scedosporium; Trichoderma reesei (Hypocreajecorina), T.citrinoviridae, T. longibrachiatum, T. virens, T. harzianum, T. asperellum, T. atroviridae,T. parareesei, Fusarium oxysporum, F. gramineanum, F. pseudograminearum, F.venenatum, Gibberella fujikuroi, G. monilformis, G. zeaea, Nectria (Haematonectria) haematococca, Stachybotrys chartarum, S. chlorohalonata, Claviceps purpurea, Metarhizium acridum, M. anisopliae, Villosiclava virens, Ophiocordyceps sinensis, Acremonium (Cephalosporium) chrysogenum, and Scedosporium apiospermum, and Aspergillus niger, Aspergillus awamori, Aspergillus oryzae, Chrysosporium lucknowense, Myceliophthora thermophila, Humicola insolens, and Humicola grisea, bacterial cells, gram-positive Bacilli such as B. subtilis, B. licheniformis, B.megaterium, B. amyloliquefaciens, B. pumilus, gram negative bacteria such as Escherichia coli, actinomycetales such as Streptomyces sp., and yeasts, such as Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, most preferably Trichoderma reesei or Bacillus.
The copending claims of Juntunen ‘424 teach GH45 (e.g. an endoglucanase); and SEQ ID NO: 1, which is a 93.3% identity match to instant SEQ ID NO: 1. Furthermore, Juntunen ‘424 teaches N210S, N215R/S, N220S and N225S substitutions, which corresponds the following substitutions with respect to instant SEQ ID NO: 1: N3S, N8R, N13S and N18S. With the recited substitutions, the SEQ ID NO: 1 linker of Juntunen ‘424 is 100% identical to instant SEQ ID NO: 1. Furthermore, SEQ ID NO: 1 of Juntunen ‘424 is a 100% identity match to residues 18-224 of instant SEQ ID NO:4, so the C-terminal residue of the cellulase in SEQ ID NO: 1 of Juntunen ‘424 is proline (relevant to instant claims 1, and 4-5). SEQ ID NO: 1 of Juntunen ‘424 is an 80.9% identity match to residues 22-228 of instant SEQ ID NO: 12 (relevant to instant claim 6). The linker component of Juntunen ‘424 is 38 residues in length, which is at least 35 amino acids (relevant to instant claim 7). Juntunen ‘424 teaches a G219S that corresponds to a G12S substitution in instant SEQ ID NO: 1 and a G244T that corresponds to a G37T substitution in instant SEQ ID NO: 1 (relevant to instant claims 9 and 27). SEQ ID NO: 1 of Juntunen ‘424 is a 100% identity match to residues 269-304 of instant SEQ ID NO: 14 (relevant to instant claim 11).
The copending claims lack: a fusion protein that has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14 (relevant to instant claim 1); a fusion protein having at least 90% identity to instant SEQ ID Nos: 16, 18 or 20 (relevant to instant claim 12); an isolated nucleic acid encoding the fusion protein (relevant to instant claim 13); the recombinant expression vector comprising the isolated nucleic acid encoding the fusion protein operably linked to regulatory sequences capable of directing expression of a gene encoding said fusion protein in a host (relevant to instant claim 14); an enzyme composition comprising the fusion protein of claim 1, and at least one additional ingredient selected from a list that includes at least one polyol selected from organic acids, lactic acid, boric acid, and proteases (relevant to instant claim 17); and a detergent thereof (relevant to instant claim 38), wherein the detergent is in the form of a liquid or solid and includes one or more additional enzymes selected from a group that includes proteases, and one or more components selected from a list that includes surfactants, builders, chelators, bleach system, dyes, perfume, and optical brighteners (relevant to instant claim 39); wherein the enzyme composition is characterized in that said organic acid is selected from a group that includes citric acid (relevant to instant claim 43). The copending claims lack the enzyme composition in the form of liquid composition or a solid (relevant to instant claim 18); wherein the enzyme composition is characterized in that said solid composition is selected from a solution, dispersion, paste, powder, granule, granulate, coated granulate, tablet, cake, crystal, crystal slurry, gel, or pellet (relevant to instant claim 44). The copending claims lack a detergent composition comprising the fusion protein, and herein the detergent composition comprises one or more component selected from a group that includes surfactants, builders, chelators or chelating agents, bleach system or bleach components, polymers, dyes, perfume, optical brighteners, anti-redepositions agents, binders, and dyes (relevant to instant claim 19); wherein the detergent is in the form of a liquid or solid and comprises one or more enzymes, such as proteases, and one or more components selected from a group that includes surfactants, builders, chelators, bleach, dyes, perfumes, and optical brighteners (relevant to instant claim 37). The copending claims lack an isolated nucleic acid encoding the fusion protein, which requires a 90% identity match to instant SEQ ID Nos: 16,18 or 20 (relevant to instant claim 28); wherein the vector is operably linked to a regulatory sequence (relevant to instant claim 29).
Chieffi teaches SEQ ID NO: 4, which is a 90.7% identity match to instant SEQ ID NO: 16 (relevant to instant claim 12). Juntunen teaches cel45A-ACM0, corresponding to nucleic acid sequence SEQ ID NO: 3. See the paragraph spanning pages 16-17 and claims 1-3 of Juntunen (relevant to instant claim 13). Juntunen teaches constructing plasmids with promoters and terminators. See page 17 first passage (relevant to instant claim 14). Juntunen teaches detergent compositions with polyols, preservatives, lactic acid, boric acid, proteases, polyethylene glycol, surfactants, builders, bleaching agents, polymers/antiredeposition agents, dyes, perfumes, and optical brighteners. See page 3 lines 23-28, and pages 12-13 and claims 16-17 of Juntunen. Cheiffi teaches detergent ingredients such as the co-builders citric acid and citrate. See paragraph [0089] (relevant to instant claims 17, 37-39 and 43). Juntunen teaches solid or liquid preparations in the form of dried powder, a tablet, a crystal or a crystal slurry on page 12 (relevant to instant claim 18 and 44). Juntunen teaches an endoglucanase polypeptide variant or an enzyme preparation thereof with surfactants, builders, bleaching agents (e.g. bleach component), polymers/antiredeposition agents, dyes, perfumes, optical brighteners, and anti-corrosion agents. See page 3 lines 23-28, and page 12 (relevant to instant claims 19 and 37). Juntunen teaches an isolated polynucleotide or complementary DNA encoding an endoglucanase polypeptide variant. See claim 11 of Juntunen. Juntunen teaches constructing expression plasmids. The constructs contain a T. reesei cel7A promoter and a terminator (e.g. regulatory sequences). See the paragraph spanning pages 16-17 (relevant to instant claims 28-29).
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention that SEQ ID NO:1 recited in copending claim 1 comprising the substitutions recited in copending claim 8 necessarily results in an at least 5-fold increase in improved protease stability compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or 14 in a liquid detergent in the present of protease (relevant to instant claim 1), and to further combine it with Chieffi’s citric acid and Juntunen’s detergent, proteases, surfactants, builders, bleaching agents, polymers/antiredeposition agents, dyes, perfumes, and optical brighteners; in order to arrive at a fusion protein that has depilling and/or antipilling effect on fabric contain cellulose or cellulose derivative; and necessarily has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein SEQ ID NO: 4 or 14 in liquid detergent in the presence of protease.
This is a provisional nonstatutory double patenting rejection.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-17 of U.S. Patent No. 10570383B2 (hereafter Juntunen ‘383) in view of Juntunen WO 2016/066896 (referred to as Juntunen), Schnorr US 9,765,373, published 09/19/2017, Payne Proceedings of the National Academy of Sciences, 2013110(36), 14646-14651, Montalibet (US 2013/0052694) and Chieffi EP 3301148 A1.
Patent claim 1 recites a cellulase variant polypeptide, or an active fragment thereof, wherein the cellulase variant polypeptide or active fragment thereof has cellulase activity and comprises an amino acid sequence having at least 90% identity to residues 22-235 of SEQ ID NO:1 comprising a substitution at one or more amino acid position(s) selected from 22 and 122, wherein at position 22 G is substituted to A, S or V and at position 122 N is substituted to A, S, G, T, V, L or I, wherein the amino acid positions of said variant polypeptide or active fragment thereof are numbered by correspondence with the mature amino acid sequence of SEQ ID NO:3.
Patent claim 7 recites an enzyme composition comprising the cellulase variant polypeptide according to claim 1.
Patent claim 8 recites the enzyme composition according to claim 7 wherein said composition further comprises other enzymes selected from the group consisting of protease, amylases, cellulases, lipases, xylanases, mannanases, cutinases, esterases, phytases, DNAses, pectinases, pectate lyases, pectinolytic enzymes, carbohydrases, arabinases, galactanases, xanthanases, xyloglucanases, laccases, peroxidases and oxidases with or without a mediator.
Patent claim 9 recites the enzyme composition according to claim 7, wherein said composition comprises a suitable additive selected from the group consisting of a stabilizer, a buffer, a surfactant, a builder, a bleaching agent, a mediator, an anti-corrosion agent, an antiredeposition agent, a caustic, an abrasive, an optical brightener, a dye, a pigment, and a preservative.
Patent claim 10 recites the enzyme composition according to claim 7, wherein said enzyme composition is in the form of a solution, dispersion, paste, powder, granule, granulate, coated granulate, tablet, cake, crystal, crystal slurry, gel, or pellet.
Patent claim 11 recites the cellulase variant polypeptide or active fragment thereof according to claim 1 for use in detergents, in treating fiber, in wood-derived pulp, in biomass hydrolysis, in food or feed application, or in any application involving modification, degradation or removal of cellulose containing material.
Patent claim 12 recites a method for antigreying, stain removal, fiber and color care, biostoning or biofinishing comprising adding the cellulase variant polypeptide according to claim 1 to liquid used in treating cotton containing fabric or garments or other textile materials.
Patent claim 13 recites the method of claim 12, wherein the textile materials are manufactured of natural cellulose containing fibers or manmade cellulose containing fibers or are mixtures thereof.
Patent claim 14 recites a detergent composition comprising the cellulase variant polypeptide or active fragment thereof according to claim 1.
Patent claim 15 of Juntunen ‘383 recites the detergent composition of claim 14 in the form of liquid detergent or a solid detergent.
Patent claim 16 recites the detergent composition according to claim 14 comprising one or more enzymes selected from the group consisting of proteases, amylases, cellulases, lipases, xylanases, mannanases, cutinases, esterases, DNAases, pectinases, pectate lyases, pectinolytic enzymes, carbohydrases, arabinases, galactanases, xanthanases, xyloglucanases, laccases, peroxidases and oxidases, preferably from the group of proteases, amylases, cellulases and lipases.
The patent claims lack: a fusion protein comprising an endoglucanase cellulase component, wherein the C-terminal amino acid of the cellulase component is changed from alanine to proline (A228P), a linker component that is at least 94% identical to SEQ ID Nos:1, 2 or 3, and, according to the numbering corresponding to the SEQ ID NO 1, comprises the following characteristics: the amino acid at position 3 is S; the amino acid at position 8 is R; the amino acid at position 13 is S; and the amino acid at position 18 is S; wherein the fusion protein has a depilling and/or antipilling effect on fabric containing cellulose or cellulose derivative; and wherein the fusion protein has at least a 5-fold increase in improved protease stability compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or 14 in a liquid detergent in the presence of a protease (relevant to instant claim 1); the cellulase component belongs to family GH45 (relevant to instant claim 4); the cellulase component has at least 80% sequence identity over the residues 18-224 with the SEQ ID NO: 4 (relevant to instant claim 5); the cellulase component has at least 80% sequence identity over the residues 22-228 with the SEQ ID NO: 12 (relevant to instant claim 6); a linker component comprises at least 35 amino acids (relevant to instant claim 7); an amino acid at positions 12 and 37 that are not G (relevant to instant claim 9); a carbohydrate binding component that has at least a 70% sequence identity with the residues 269-304 of SEQ ID NO: 14 (relevant to instant claim 11); a fusion protein having at least 90% sequence identity with SEQ ID NO: 16, 18 or 20 (relevant to instant claim 12). The patent claims an isolated nucleic acid encoding the fusion protein(relevant to instant claim 13). The patent claims lack a recombinant expression vector comprising a regulatory sequence capable of directing expression of a gene encoding said fusion protein in a host (relevant to instant claim 14). The patent claims lack an amino acid position at position 12 that is S and at position 37 is T (relevant to instant claim 27); an isolated nucleic acid encoding a fusion protein that is 90% identity match to instant SEQ ID Nos: 16,18 or 20 (relevant to instant claim 28); wherein the vector is operably linked to a regulatory sequence (relevant to instant claim 29). The patent claims of Juntunen ‘383 lack an enzyme composition characterized in that said organic acid is selected from a group that includes citric acid (relevant to instant claim 43)
However, Juntunen teaches a cel45A-ACM0 corresponding to SEQ ID NO: 12 containing a catalytic core attached to a linker and a carbohydrate binding module (CBM) region. See the sentence spanning p. 16-17. Juntunen’s SEQ ID NO: 12 and Chieffi’s SEQ ID NO: 4 includes a P residue corresponding to the C-terminal residue 224 of instant SEQ ID NO: 4. Juntunen discloses that cellulases are used to prevent formation of pills (e.g. antipilling) on the surface of cotton garments (e.g. fabric containing cellulose). See p. 1 lines 27-28. Schnorr teaches common substitutions, such as Ser/Asn. See column 16 lines 8-10. Payne suggests that serine residues often exhibit O-glycosylation, which imparts protease resistance. See the first two paragraphs on page 14646. Montalibet teaches an N431R substitution. See claim 9 of Montalibet (relevant to instant claim 1). Juntunen teaches variants endoglucanase peptides belonging to the glycosyl hydrolase family 45, i.e. GH45. See the first full paragraph on page 6 (relevant to instant claim 4). Juntunen teaches SEQ ID NO: 12, which is a 96.7% identity match over residues 18-224 of instant SEQ ID NO: 4; an 80.9% identity match over residues 22-228 of instant SEQ ID NO: 12; and the linker in SEQ ID NO: 12 is at least 35 amino acids in length. See the sequence alignment attached in the office action appendix of the non-final action mailed 07/09/2024 (relevant to instant claims 5-7). Schnorr suggests that N may be replaced with S, and G may be replaced with S or T within proteins having cellulolytic activity. See columns 15-16. Payne teaches that serine and threonine residues in linkers may impart protease resistance. See the first two paragraphs on page 14646 of Payne (relevant to instant claims 9 and 26-27). Juntunen teaches SEQ ID NO: 12, which is a 100% identity match over residues 269-304 of instant SEQ ID NO: 14 (relevant to instant claim 11). Chieffi teaches SEQ ID NO: 4, which is a 90.7% identity match to instant SEQ ID NO: 16; and includes a P residue at position 224 relative to the cellulase component of instant SEQ ID NO: 4 (relevant to instant claim 12). Juntunen teaches cel45A-ACM0, corresponding to nucleic acid sequence SEQ ID NO: 3. The nucleic acid sequence may include a promoter and a terminator, such that the polynucleotide is operably linked to regulatory sequences. See the paragraph spanning pages 16-17 and claims 1-3 and 10-12 of Juntunen (relevant to instant claims 13-14 and 28-29). Cheiffi teaches detergent ingredients such as the co-builders citric acid and citrate. See paragraph [0089] (relevant to instant claim 43).
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to replace the cellulase variant polypeptide recited in patent claim 1 with the cellulase and CBM domains of SEQ ID NO: 4 of Chieffi and the linker of Juntunen and to further modify the linker region of Juntunen in view of Schnorr, Payne and Montalibet in order to arrive at a fusion protein that has depilling and/or antipilling effect on fabric contain cellulose or cellulose derivative; and necessarily has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein SEQ ID NO: 4 or 14 in liquid detergent in the presence of protease.
Claims 1, 4-7, 9, 11-14, 17-19, 27-29, 37-39 and 43-44 are rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-21 of U.S. Patent No. 9994836 B2 (hereafter Juntunen ‘836) in view of Juntunen WO 2016/066896 (referred to as Juntunen), Schnorr US 9,765,373, published 09/19/2017, Payne Proceedings of the National Academy of Sciences, 2013110(36), 14646-14651, Montalibet (US 2013/0052694), and Chieffi EP 3301148 A1. The underlined text below is relevant to the instant claims. The conflicting patent US9994836 is in the same family as Juntunen WO 2016/066896, so the sequence alignments are the same.
Patent claim 1 recites a variant endoglucanase polypeptide comprising an amino acid sequence having at least 95% sequence identity with amino acids 1 to 214 of SEQ ID NO: 1 and an amino acid substitution at three or more positions selected from the group consisting of 51, 75, 77, 82, 109, 116, 118, 135, 150 and 205, the positions being numbered with reference to SEQ ID NO: 1, wherein the variant polypeptide has endoglucanase activity.
Patent claim 2 of Juntunen ‘836 recites the variant endoglucanase polypeptide of claim 1 attached to a carbohydrate binding module and optionally to a linker region.
Patent claim 3 of Juntunen ‘836 recites the variant endoglucanase polypeptide of claim 1, wherein the variant endoglucanase polypeptide is attached to a carbohydrate binding module and a linker region, which are heterologous or homologous.
Patent claim 4 of Juntunen ‘836 recites the variant endoglucanase polypeptide of claim 3, wherein at least one of the carbohydrate binding module and the linker region is heterologous.
Patent claim 8 of Juntunen ‘836 recites the variant endoglucanase polypeptide of claim 1, wherein the polypeptide has an amino acid sequence of SEQ ID NO: 12 and an amino acid substitution at three or more positions selected from the group consisting of 51, 75, 77, 82, 109, 116, 118, 135, 150 and 205, the positions being numbered with reference to SEQ ID NO: 1.
Patent claim 11 of Juntunen ‘836 recites an isolated polynucleotide comprising: a) a polynucleotide or complementary DNA encoding an endoglucanase polypeptide variant of claim 1; b) a polynucleotide encoding a fragment of a polypeptide encoded by a polynucleotide of a) wherein said fragment is having endoglucanase activity, c) a polynucleotide comprising a nucleotide sequence which is degenerate to the nucleotide sequence of a polynucleotide sequence of a) or b), or the complementary strand of such a polynucleotide.
Patent claim 12 of Juntunen ‘836 recites a vector, which comprises an isolated polynucleotide of claim 11 operably linked to regulatory sequences capable of directing expression of the endoglucanase polypeptide variant.
Patent claim 14 of Juntunen ‘836 recites a method of producing the endoglucanase polypeptide variant of claim 1, said method comprising the steps of transforming a host cell with an expression vector encoding said polypeptide, and culturing said host cell under conditions enabling expression of said polypeptide, and optionally recovering and purifying said polypeptide variant.
Patent claim 15 of Juntunen ‘836 recites an enzyme preparation comprising the endoglucanase polypeptide variant according to claim 1.
Patent claim 16 of Juntunen ‘836 recites the enzyme preparation of claim 15 further comprising at least one enzyme selected from the group consisting of other cellulases, amylases, lipases, proteases, hemicellulases, ligninases, pectinolytic enzymes and oxidative enzymes.
Patent claim 17 of Juntunen ‘836 recites a detergent composition comprising the endoglucanase polypeptide variant of claim 1 or an enzyme preparation thereof, a surfactant and optionally one or more additives selected from the group consisting of stabilizers, buffers, surface active agents, builders, cobuilders, bleaching agents, bleach activators, other detergent enzymes, mediators of oxidative enzymes, anti-corrosion agents, antiredeposition agents and soil release polymers, caustics, abrasives, optical brighteners, dyes, pigments, perfumes and preservatives.
Patent claim 20 of Juntunen ‘836 recites the method of claim 19, wherein the endoglucanase polypeptide variant is within a detergent composition and the detergent composition is contacted to the textile material.
The patent claims of Juntunen ‘836 teach a variant endoglucanase polypeptide (e.g. a fusion peptide) comprising a carbohydrate binding module and a linker region, including SEQ ID NO: 12, which contains a linker region, residues 225-262, that is 93.3% identical to instant SEQ ID NO: 1; however the linker of Juntunen ‘836 is not 94% identical to instant SEQ ID NO: 1 (relevant to instant claim 1). The patent claims of Juntunen ‘836 teach SEQ ID NO: 12, which is a 96.7% identity match over residues 18-224 of instant SEQ ID NO: 4 (relevant to instant claim 5), and an 80.9% identity match over residues 22-228 of instant SEQ ID NO: 12 (relevant to instant claim 6). Juntunen ‘836 teach a linker region that is at least 35 amino acids in length (relevant to instant claim 7). See the alignment provided in the appendix of the non-final action mailed 07/09/2024. SEQ ID NO: 12 in the patent claims of Juntunen ‘836 is a 100% identity match over residues 269-304 of instant SEQ ID NO: 14 (relevant to instant claim 11).
The patent claims lack: a cellulose component wherein the C-terminal amino acid is changed from alanine to proline (A228P), a linker component that is at least 94% identical to SEQ ID Nos:1, 2 or 3, and, according to the numbering corresponding to the SEQ ID NO 1, comprises the following characteristics: the amino acid at position 3 is S; the amino acid at position 8 is R; the amino acid at position 13 is S; and the amino acid at position 18 is S; wherein the fusion protein has a depilling and/or antipilling effect on fabric containing cellulose or cellulose derivative; and wherein the fusion protein has at least a 5-fold increase in improved protease stability compared to a reference fusion protein having the amino acid sequence SEQ ID NO: 4 or SEQ ID NO: 14 in a liquid detergent in the presence of a protease (relevant to instant claim 1); the cellulase component belongs to family GH45 (relevant to instant claim 4); amino acid position at positions 12 and 37 that are not G (relevant to instant claim 9); fusion protein having at least 90% sequence identity with SEQ ID NO: 16, 18 or 20 (relevant to instant claim 12); an enzyme composition in the form of a solution, dispersion, paste, powder, granule, granulate, coated granulate, tablet, cake, crystal, crystal slurry, gel, or pellet (relevant to instant claim 18 and 44); amino acid position at position 12 that is S and at position 37 is T (relevant to instant claim 27); an isolated nucleic acid encoding the fusion protein of instant claim 12, which requires a 90% identity match to instant SEQ ID Nos: 16,18 or 20 (relevant to instant claim 28); wherein the vector is operably linked to a regulatory sequence (relevant to instant claim 29); a detergent composition in the form of liquid or solid (relevant to instant claim 39); an enzyme composition characterized in that said organic acid is selected from a list that includes citric acid (relevant to instant claim 43).
However, Juntunen teaches a cel45A-ACM0 corresponding to SEQ ID NO: 12 containing a catalytic core attached to a linker and a carbohydrate binding module (CBM) region. See the sentence spanning p. 16-17. Juntunen’s SEQ ID NO: 12 and Chieffi’s SEQ ID NO: 4 includes a P residue corresponding to the C-terminal residue 224 of instant SEQ ID NO: 4. Juntunen discloses that cellulases are used to prevent formation of pills (e.g. antipilling) on the surface of cotton garments (e.g. fabric containing cellulose). See p. 1 lines 27-28. Schnorr teaches common substitutions, such as Ser/Asn. See column 16 lines 8-10. Payne suggests that serine residues often exhibit O-glycosylation, which imparts protease resistance. See the first two paragraphs on page 14646. Montalibet teaches an N431R substitution. See claim 9 of Montalibet (relevant to instant claim 1). Juntunen teaches variants endoglucanase peptides belonging to the glycosyl hydrolase family 45, i.e. GH45. See the first full paragraph on page 6 (relevant to instant claim 4). Chieffi teaches SEQ ID NO: 4, which is a 90.7% identity match to instant SEQ ID NO: 16. See the alignments provided in the office action appendix of the non-final action mailed 07/09/2024 and above, and see lines 3-4 on page 9 of Juntunen (relevant to instant claim 12). Schnorr suggests that G may be replaced with S or T within proteins having cellulolytic activity. See columns 15-16 (relevant to instant claim 27). Juntunen teaches cel45A-ACM0, corresponding to nucleic acid sequence SEQ ID NO: 3. The nucleic acid sequence may include a promoter and a terminator, such that the polynucleotide is operably linked to regulatory sequences. See the paragraph spanning pages 16-17 and claims 1-3 and 10-12 of Juntunen (relevant to instant claims 28-29). Chieffi teaches detergent ingredients such as the co-builders citric acid and citrate. See paragraph [0089] (relevant to instant claim 43).
It would have been obvious to a person of ordinary skill in the art prior to the effective filing date of the instantly claimed invention to replace endoglucanase variant polypeptide recited in patent claim 1 with the cellulase and CBM domains of SEQ ID NO: 4 of Chieffi and the linker of Juntunen and to further modify the linker region of Juntunen in view of Schnorr, Payne and Montalibet in order to arrive at a fusion protein that has depilling and/or antipilling effect on fabric contain cellulose or cellulose derivative; and necessarily has at least a 5-fold increase in improved protease stability, compared to a reference fusion protein SEQ ID NO: 4 or 14 in liquid detergent in the presence of protease.
Response to Arguments
Applicant's arguments filed 04/24/2026 have been fully considered but they are unpersuasive.
Double Patenting Rejections
Applicant argues that the cited art fails to teach or suggest the specific positional substitution required by claim 1, which are shown in the specification to provide improved and surprising technical effect (see example 2 and figures 2 and 3). Specifically, claim 1 is structurally and functionally distinct from the claims of the cited references. See the remarks the paragraph spanning pages 14-15. Applicant argues that the A228P mutation is not cited in the prior Juntunen Patents. This provides clear structural distinction from the cited patents. Juntunen ‘424 focusses on specific substitutions within catalytic core and does not require this mutation. Juntunen ‘383 and Juntunen ‘836 do not require the A228P mutation. See the remarks p. 14 section 1. Applicant argues that the claims recite a patentably distinct engineered motif. The instantly claimed motif consists of the A228P junction and the serine and arginine substitutions in the linker. As recited, this structural combination yields a 5-fold increase in improved protease stability. See the remarks p. 14 section 2.
This argument is not persuasive because the results relied upon are not commensurate in scope with the instant claims. Example 2 teaches testing the color revival of MA79+CBM and MA79+CBML. See p. 28 lines 29-30. MA79+CBM is SEQ ID NO: 14. See p. 6 line 26. MA79+CBML is SEQ ID NO: 18. See p. 7 line 9. The specification teaches that the C-terminal amino acid of the cellulase component is Pro, corresponding to the substitution A228P in SEQ ID NO: 14. See p. 22 lines 12-13. SEQ ID NOs 14 and 18 include a P228 residue. Figures 2-3 show that MA79+CBML (SEQ ID NO: 18) has increased darkness results in terms of color revival compared to MA79+CBM. However, the instant claims do not limit the fusion protein to MA79+CBML (SEQ ID NO: 18), nor does the specification clearly attribute increased darkness to the P228 residue because P228 is present in SEQ ID NO: 14. Furthermore, Applicant argues that copending application Juntunen ‘424 does not teach the A228P residue. Claim 1 does not clearly require any specific A228P substitution because the claim does limit the cellulase sequence, so it is unclear which position 228 is being referenced in the claim. The specification discloses that SEQ ID NO: 4 (residues 18-224) is a cellulase component. SEQ ID NO: 1 of Juntunen ‘424 is a 100% identity match to residues 18-224 of instant SEQ ID NO:4. Therefore, SEQ ID NO: 1 recited in the copending claims of Juntunen ‘424 has the required C-terminal proline. Furthermore, Juntunen ‘383 and ‘836 are not relied upon for teaching the cellulase component.
Conclusion
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/LOUISE W HUMPHREY/Supervisory Patent Examiner, Art Unit 1657
/K.C.B./Examiner, Art Unit 1657
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