DETAILED ACTION
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 .
Status of Claims
Claims 1-3, 6 and 9 are pending following the Reply filed 09/11/2025. Claims 1-3 and 9 have been amended without introducing new matter. Claims 4-5, 7-8 and 10 have been cancelled. Claims 1-3, 6 and 9 have been examined on the merits.
Information Disclosure Statement
The information disclosure statement (IDS) filed on 11/11/2025 is in compliance with the provisions of 37 CFR 1.97 and has been considered by the examiner.
Withdrawn
Any objection or rejection of claims 4-5, 7-8 and 10 is moot, because the claims have been cancelled.
The objections of claims 1 and 3 are withdrawn in light of the amendments.
The rejections of claims 2-3 under 35 U.S.C. 112(b) are withdrawn in light of the amendments.
The rejection of claims 1-3, 6 and 9 under 35 U.S.C. 103 has been withdrawn in light of the declaration filed 09/11/2025 and Applicant’s Arguments. See Response to Arguments and Allowable Subject Matter below for further discussion.
Claim objections
Claim 1 is objected to because of the following informalities: please amend “derived from Genus Shewanella microorganism” in lines 5-6 to “derived from a Genus Shewanella microorganism. Appropriate correction is required.
Maintained Rejections and New Rejections Necessitated by Amendment
Claim Rejections - 35 USC § 112(a) – Scope of Enablement
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 1-3, 6 and 9 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for a Genus Corynebacterium microorganism expressing SEQ ID NO: 1, does not reasonably provide enablement for the microorganism expressing any membrane protein having at least 75% sequence identity to SEQ ID NO: 1. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the invention commensurate in scope with these claims.
Enablement requires that the specification teach those in the art to make and use the
invention without undue experimentation. Factors to be considered in determining whether a
disclosure would require undue experimentation include (1) the nature of the invention, (2) the state of the prior art, (3) the predictability or lack thereof in the art, (4) the amount of direction or guidance present, (5) the presence or absence of working examples, (6) the quantity of experimentation necessary, (7) the relative skill of those in the art, and (8) the breadth of the claims.
The nature of the invention – Claim 1 recites “wherein the membrane protein is derived from [a] Genus Shewanella microorganism”. This limitation does not meaningfully limit the structure of the claimed product, and the claims are directed to ANY membrane protein having at least 75% sequence identity to SEQ ID NO: 1. A person having ordinary skill in the art would recognize this genus to include many undisclosed structures, including those that have been structurally modified from the wildtype protein derived from Shewanella. Thus, the claimed genus includes variants of the membrane protein having as many as 50 amino acid modifications relative to SEQ ID NO: 1. Similarly, claim 3 recites the “nucleic acid sequence having a sequence identity of at least 75%”, which would include variants with as many as 152 base pair modifications.
Hence, the full scope of the claims encompasses ANY variant of the claimed protein having at least 75% sequence identity to SEQ ID NO: 1, wherein the microorganism expressing the protein, or variant thereof, has increased productivity in the production of L-lysine or L-arginine. As further discussed below, the limitations requiring the membrane protein, and/or the nucleic acid encoding it, to have a sequence identity of at least 75%, to SEQ ID NOs 1 and 2 respectively, still includes what are potentially trillions of variants.
The state of the prior art – A review by Becker et al. published in 2012 (previously cited; hereafter “Becker”) discusses recent advances in systems and synthetic metabolic engineering for amino acids and related compounds, covering major achievements for the two major industrial production organisms, Corynebacterium glutamicum and Escherichia coli (see pg. 718, col. 2, para. 1). Becker teaches that the development of over-producing strains in amino acid production has been carried out for many decades by iterative rounds of random mutagenesis to provide mutants with remarkable performance, that were however, limited by growth defects, low stress tolerance or enhanced nutrient demand owing to the inherent accumulation of detrimental mutations during strain development (see pg. 718, col. 2, para. 2). Becker teaches that in the shift to rational approaches, enabled by recombinant DNA technology, it has not been easy to reach the high performance of classical strains for existing industrial processes (see pg. 718, col. 2, para. 2). Becker teaches this particularly holds true for amino acid production, which demands a global modification of pathway fluxes within a complex interconnected network of reactions that is tightly constrained by different layers of metabolic and transcriptional regulation (see pg. 718, col. 2, para. 2). In conclusion, Becker states that with the exception of the most advanced example of lysine production, the production properties of rationally created amino acid producers created so far still leaves space for substantial improvement in the future (see pg. 723, col. 2, para. 2).
The predictability or lack thereof in the art -- Whisstock, et al. (cited in the IDS filed 02/16/2024) teach that the prediction of a protein’s function based on its protein sequence is difficult, because homologous proteins often have different functions (see Abstract; pg. 323, para. 1). Whisstock et al. also teach that although it is possible homologous proteins will share similar functions, the substitution of a single, critically placed amino acid in an active site residue may be sufficient to alter a protein’s role fundamentally (see pg. 323, para. 1). Witkowski et al. (cited in the IDS filed 02/16/2024) teach that a single conservative replacement of an amino acid transforms β-ketoacyl synthase to malonyl decarboxylase and completely eliminates β-ketoacyl synthase activity (see Abstract, Table 1). Seffernick et al. (cited in the IDS filed 02/16/2024) teach that two enzymes derived from Pseudomonas sp., sharing 98% amino acid sequence identity, catalyze two different reactions (i.e., deamination versus dehalogenation) and, hence, possess different functions (see Abstract). Hence, in view of the prior art of record, it would appear that even a slight modification to an existing protein may have unpredictable effects on its function, and this unpredictability would reasonably be expected to increase with the number of modifications being made.
The amount of direction or guidance present – The present inventors disclose that a foreign L-amino acid exporting protein with high L-amino acid exporting activity was searched, and by introducing this to a lysine producing strain, a recombinant strain with improved L-amino acid exporting ability and/or productivity was provided (see, e.g., pg. 3, lines 2-5). According to the specification, the amino acid sequence of the foreign protein or the nucleic acid sequence encoding it may be modified by common mutagenesis “within the range of maintaining its original function and/or desired function” (see pg. 8, lines 2-5). Applicant discloses methods and systems known in the art for determining the homology, similarity or identity of polynucleotide and polypeptide sequences, such as, computer algorithms, programs and database webtools (see pgs. 8-10), but there is no guidance in the disclosure as to what structure(s) in the claimed sequence correspond to the desired functions of the membrane protein, its derivatives or variants.
The presence or absence of working examples – In Applicant’s Examples, strains of Corynebacterium glutamicum were transformed by the introduction of a membrane protein derived from Shewanella atlantica having the amino acid sequence of SEQ ID NO: 1 (see pg. 18, lines 10-12; pg. 20, lines 6-7). The Examples of Applicant’s disclosure do not disclose any working examples using a microorganism expressing a variant of this amino acid sequence or an amino acid sequence derived from another species of Shewanella. Even if the membrane protein used in Applicant’s Examples were modified in some way relative to its parent sequence, it is not disclosed on record how or to what extent it differed from the recited sequence. Hence, there are no working examples present in the specification comprising variants of SEQ ID NO: 1 or using membrane proteins derived from other members of Shewanella.
The quantity of experimentation necessary – As no single definition has been set forth on record of what constitutes a “derivative”, one of ordinary skill could not envision any and all means to derive variants from Shewanella membrane proteins that have improved productivity, as is necessary to practice the full scope of the claimed invention. Furthermore, the genus of variant membrane proteins comprising at least 75% identity with SEQ ID NO: 1 requires the modifying of up to 50 amino acid residues in a sequence comprising 202 amino acids. In view of Reetz, MT. (previously cited), the substitution of only three amino acid residues in a polypeptide composed of 300 residues can result in about 30 billion variants (see pg. 140, col. 1, para. 2 to col. 2, para. 1). Moreover, there is no prior art teaching or guidance provided in the present disclosure as to how mutating SEQ ID NO: 1 would result in a functioning membrane protein, let alone one that would enable the bacterium to have increased productivity of L-lysine or L-arginine.
Hence, as there are no explicit teachings in the prior art or guidance found in the present disclosure to determine which consensus structure(s) correspond to the claimed function(s) of the membrane protein derived from SEQ ID NO: 1, a person of ordinary skill would be required to experiment with billions of variants in order to practice the full scope of the claimed invention.
The relative skill of those in the art – While the level of skill in the art is high, predictability in the art is low due to the complexity of metabolic and transcriptional pathways, protein structure/function relationships, and the response of host strains to biological engineering. Accordingly, the skilled artisan would not be able to envisage, let alone practice, the full scope of the claimed invention given the breadth of the claims and the lack of guidance in the specification.
The breadth of the claims – In order to make and use the full scope of the invention, as set forth above, the claims require the engineering of membrane protein variants, out of potentially trillions of variants, that not only retain the function of a membrane protein, but also impart the enhanced function of increased productivity of L-lysine or L-arginine when introduced into a Corynebacterium.
Accordingly, the quantity of experimentation required to determine which derivatives/variants of the membrane protein can perform these functions constitutes an invitation to experiment without any reasonable expectation of success even given the relative skill of those in the art.
Response to Arguments
Regarding the rejections under 35 U.S.C. 112(a) – Scope of Enablement, Applicant argues that claim 1 has been “tightened”, because: (1) the claim now anchors the membrane protein to SEQ ID NO: 1 itself or to sequences with at least 75% identity to that specific sequence, while requiring that the protein be derived from genus Shewanella; and (2) the claim is functionally limited to the very result demonstrated and taught in the specification of increased productivity of L-lysine or L-arginine in Corynebacterium strains.
Applicant’s arguments have been fully considered but are not persuasive for the following reasons.
First, in the claims filed 09/30/2022, claim 1 recited “a membrane protein derived from Shewanella atlantica of SEQ ID NO: 1 or a membrane protein having a sequence homology of at least 75% or more thereto”, whereas amended claim 1 now recites “wherein the membrane protein is derived from [a] Genus Shewanella microorganism”. While the membrane protein having at least 75% sequence identity to SEQ ID NO: 1 is now required to be derived from a genus Shewanella microorganism, this limitation does not limit the structure of the end-product, which still includes variants of the membrane protein that have not been disclosed.
Second, while the claims have been narrowed to “increased productivity of L-lysine or L-arginine” rather than the increased productivity of any L-amino acid, this functional limitation must be enabled for the full scope of the claimed genus, which includes trillions of potential species. This is similar to Amgen Inc. v. Sanofi, Aventisub LLC, 987 F.3d 1080 (Fed. Cir. 2021), as affirmed by the Supreme Court, wherein the Federal Circuit relied on evidence showing that the scope of the claims encompassed millions of antibodies and that it was necessary to screen each candidate antibody in order to determine whether it met the functional limitations of the claim. Id. at 1088. Consequently, the Federal Circuit concluded that there was a lack of enablement. See MPEP 2164.01(a) and 2164.06(II). In the instant case, a person of ordinary skill would be required to screen at least millions of candidate membrane proteins in order to find ones that meet the functional limitation.
Applicant further argues that the “≥75% identity” space around SEQ ID NO: 1 is a compact, genus-restricted set of LysE/ArgO family transporters from Shewanella. The BLAST output submitted of record shows that all of the top hits at identities well above the claimed 75% threshold are LysE/ArgO family transporters from Shewanella species, with identities ranging from approximately 98% down to the high 70s, all annotated within the same transporter family and taxonomic genus. This demonstrates that the claim's identity-based alternative is not an open-ended invitation to explore "trillions of variants," but rather a well-defined and predictable sequence set that a skilled person can identify using ordinary tools exactly as taught in the specification.
Applicant’s arguments have been fully considered but are not persuasive for the following reasons.
First, the only working example in Applicant’s disclosure was directed to a membrane protein derived from Shewanella atlantica with 100% sequence identity to SEQ ID NO: 1. No other species alluded to in Applicant’s argument (i.e., proteins derived from Shewanella canadensis, Shewanella sediminis, or Shewanella benthica) or any possessing 75-98% sequence identity were reduced to practice or even discussed in Applicant’s disclosure. Applicant acknowledges that the claimed results were “unexpected” (see Remarks at pg. 7) which is evidenced by the Rule 1.132 Declaration filed on 09/11/2025. Hence, there is no evidence or reasonable expectation that any LysE/ArgO family protein derived from the same bacterial genus would give the same results as the membrane protein of Applicant’s disclosure, particularly when these results are alleged to have been unpredictable.
Furthermore, the membrane protein having at least 75% sequence identity to SEQ ID NO: 1 is reasonably interpreted to be the end-product of this limitation, and the species that this protein is “derived from” does not limit its structure to only those that are found in public databases. As discussed under the present rejection, there is no language in the claim that would exclude membrane proteins derived from Shewanella to be genetically modified variants, and this genus therefore comprises trillions of potential variants.
Regarding the rejections under 35 U.S.C. 103, Applicant argues that the proposed prior art combination lacks a reasoned motivation or reasonable expectation of success because: (1) Ueda neither identifies or suggests Shewanella LysE/ArgO family sequences; and (2) the GenBank record for RTR33311.1 is a bare sequence deposit and does not teach expression in Corynebacterium or any fermentation-productivity benefit. The data in the Declaration also confirm that the claimed results were unexpected and underscore the host-transporter specificity and unpredictability of exporter/host interactions that the instant specification and the broader literature describe.
Applicant’s arguments have been fully considered and are persuasive. Accordingly, the rejection of claims 1-3, 6 and 9 under 35 U.S.C. 103 has been withdrawn. See Allowable Subject Matter below for further discussion.
Allowable Subject Matter
The declaration under 37 CFR 1.132 filed 09/11/2025 is sufficient to overcome the rejection of claims 1-3, 6 and 9 under 35 U.S.C. 103 based upon the Ueda and GenBank RTR33311.1 references, because the declaration provides experimental evidence demonstrating that not all candidate L-lysine export membrane proteins enhance L-amino acid production. In addition to the factors discussed below, the evidence provided in the Declaration further demonstrates that selecting the claimed membrane protein for the increased productivity of L-lysine or L-arginine in Corynebacterium would have been unpredictable at the time of filing. Accordingly, it would not have been obvious for a person of ordinary skill to have selected, for example, any GenBank sequence denoted as an “amino acid exporter” or “lysine/arginine exporter” for use in Ueda’s methods with a reasonable expectation of success, and the results of Applicant’s disclosure are considered to be unexpected in view of the filed Declaration and the closest prior art of record:
Ueda (previously cited) teaches that L-amino acids are industrially produced by fermentation using microorganisms belonging to the genus Corynebacterium (see pg. 1, para. 2). Ueda teaches it is desirable to obtain a gene for an L-amino acid-exporter that exhibits an ability to secrete sufficient amounts of L-amino acids, including L-lysine and L-arginine, in a variety of heterogeneous host microorganisms (see pg. 3, para. 1). Ueda discloses a novel gene for an L-amino acid exporter, and a parent strain, which is a Coryneform bacterium, wherein the parent strain may not inherently possess the L-amino acid exporter gene but exhibits improved L-amino acid-export ability when the gene is introduced (see pg. 5, paras. 4 and 9; pg. 8, para. 3). Ueda teaches that the novel gene was predicted to encode a membrane protein and be involved in amino acid export (see pg. 43, para. 5). Ueda teaches the Coryneform bacteria to include those in the genus Corynebacterium (see pg. 10, para. 2).
However, Ueda does not teach nor suggest the L-amino acid exporter to be derived from Shewanella or to have at least 75% sequence identity to instant SEQ ID NO: 1.
GenBank RTR33311.1 (previously cited) is identified as an “amino acid transporter” derived from Shewanella atlantica strain HAW-EB5 isolated from Atlantic Ocean sediment (see Definition, Organism, and Source). The amino acid sequence is further identified as an arginine exporter protein (see Region). As discussed in the previous office action, instant SEQ ID NO: 1 is identical to RTR33311.1 and it would have been obvious for a person of ordinary skill to have selected any “amino acid transporter” or “lysine/arginine exporter” under the “simple substitution” rationale of MPEP 2143(I).
However, upon reconsideration, the examiner notes that Ueda does not teach or suggest expressing an L-amino acid exporter from the Shewanella genus, or one having any similar structural identity to the claimed protein, in a Corynebacterium. Per MPEP 2144.08, determining whether one of ordinary skill in the art would have had a reason to select the claimed species or subgenus (when the prior art teaches genus) includes considering the size of the genus, express teachings, teachings of similar properties or uses, and predictability of the technology. In the instant case, the prior art combination required selecting the claimed species from a broad genus (i.e., all L-amino acid exporters), and the examiner has not found any relevant teachings that would have led one to select the claimed subgenus (i.e., the microorganism expressing a membrane protein having at least 75% sequence identity to SEQ ID NO: 1), and particularly not the species exemplified by the disclosure (i.e., the microorganism expressing SEQ ID NO: 1), for improving L-lysine or L-arginine productivity. Moreover, the examiner has not found any relevant teachings that would have been sufficient to overcome the unpredictability of selecting such a protein from this genus for use in the claimed invention.
After an updated search, there is no teaching or suggestion in the prior art of record to render obvious a microorganism of the genus Corynebacterium expressing a Shewanella membrane protein having at least 75% sequence identity to SEQ ID NO: 1 with increased productivity of L-lysine or L-arginine.
In the absence of evidence to the contrary, which is the burden of the Patent Office to produce, claims 1-3, 6 and 9 are considered to be free of the prior art of record and are nonobvious in view of the closest prior art.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/DENNIS IGNATIUS ARMATO JR/Examiner, Art Unit 1651
/MELENIE L GORDON/Supervisory Patent Examiner, Art Unit 1651
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