tyrB

UniProt ID: Q88LG1
Organism: Pseudomonas putida (strain ATCC 47054 / DSM 6125 / CFBP 8728 / NCIMB 11950 / KT2440)
Review Status: DRAFT
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Gene Description

tyrB (PP_1972; also referred to as tyrB-1) is a cytoplasmic, pyridoxal 5'-phosphate (PLP)-dependent aminotransferase of the class-I (fold-type I) aspartate aminotransferase superfamily. It catalyzes reversible transamination in which an amino group is transferred from an amino acid donor to a 2-oxoacid acceptor (commonly 2-oxoglutarate, yielding L-glutamate). It is annotated as an aromatic-amino-acid aminotransferase, interconverting aromatic amino acids (L-tyrosine, L-phenylalanine) and their cognate aromatic 2-oxoacids (4-hydroxyphenylpyruvate, phenylpyruvate), and contributes to aromatic amino acid biosynthesis and catabolism. Like many class-I PLP aminotransferases it is a homodimer with active sites formed at the subunit interface. In P. putida KT2440 it is one of several aminotransferase isozymes acting on aromatic amino acids; genetic studies show that loss of tyrB alone causes only mild aromatic-amino-acid utilization phenotypes because of redundancy with paralogous aminotransferases (notably PP_3590/AmaC and tyrB-2/phhC), so its physiological role overlaps with those enzymes.

Existing Annotations Review

GO Term Evidence Action Reason
GO:0003824 catalytic activity
IEA
GO_REF:0000002 		MARK AS OVER ANNOTATED
Summary: Generic root-level molecular function term. tyrB is an enzyme, so this is not wrong, but it is uninformatively general and is fully subsumed by the more specific transaminase/aminotransferase activity terms.
Reason: "catalytic activity" is the MF root and conveys no specific information. The more precise terms GO:0008483 (transaminase activity) and GO:0004838 (L-tyrosine:2-oxoglutarate transaminase activity) capture the actual function.
GO:0004838 L-tyrosine:2-oxoglutarate transaminase activity
IEA
GO_REF:0000118 		ACCEPT
Summary: Specific aromatic (tyrosine) aminotransferase activity assigned by TreeGrafter from the PTHR11879:SF37 "aromatic-amino-acid aminotransferase" subfamily. This is consistent with the protein family, the COG1448 assignment, and with biochemical/genetic characterization of P. putida aromatic aminotransferases. This is the best representation of the gene's core molecular function.
Reason: Domain/family evidence (class-I PLP aminotransferase, PANTHER ArAT subfamily SF37) plus genetic evidence in KT2440 (tyrB mutants show a tyrosine-utilization phenotype; PMID:20050871) support tyrosine aminotransferase activity. The enzyme is likely promiscuous across aromatic amino acids (also acting on phenylalanine), but this term well captures the central characterized activity.
GO:0005829 cytosol
IEA
GO_REF:0000118 		ACCEPT
Summary: Cytosolic localization predicted by TreeGrafter. Soluble class-I PLP aminotransferases acting in amino acid metabolism are cytoplasmic enzymes; the sequence has no signal peptide or transmembrane region. Consistent with the expected localization.
Reason: Aromatic aminotransferases in this family are soluble cytoplasmic enzymes. No experimental KT2440 localization data exist, but the prediction is biologically appropriate and there is no evidence for periplasmic/membrane localization.
GO:0006520 amino acid metabolic process
IEA
GO_REF:0000002 		MODIFY
Summary: Broad biological process term. tyrB participates in (aromatic) amino acid metabolism, so this is correct but general. A more specific process such as aromatic amino acid family metabolism / phenylalanine or tyrosine biosynthesis or catabolism would be more informative.
Reason: The annotation is correct in essence but too high-level. tyrB acts specifically on aromatic amino acids (Tyr/Phe), so the more specific "aromatic amino acid family metabolic process" better reflects the characterized role while remaining defensible from family + genetic evidence. (Chorismate metabolic process is not appropriate: tyrB acts downstream of chorismate on the aromatic amino acids/2-oxoacids, not on chorismate itself.)
GO:0008483 transaminase activity
IEA
GO_REF:0000120 		KEEP AS NON CORE
Summary: General transaminase (aminotransferase) activity. Correct and well supported by the class-I PLP-dependent aminotransferase family assignment, but less specific than GO:0004838. Useful as a parent term.
Reason: Accurately describes the enzymatic class but is a parent of the more specific aromatic aminotransferase term that represents the core function. Retain as supporting/non-core rather than as the primary MF.
GO:0030170 pyridoxal phosphate binding
IEA
GO_REF:0000120 		ACCEPT
Summary: PLP cofactor binding. tyrB is a PLP-dependent enzyme (UniProt COFACTOR: pyridoxal 5'-phosphate; conserved PROSITE PS00105 class-I aminotransferase PLP-binding motif). This is a well-supported and informative molecular function annotation.
Reason: Strong family/motif evidence (IPR004838/IPR004839, PROSITE AA_TRANSFER_ CLASS_1, UniProt cofactor annotation) for PLP binding, which is essential for the transamination mechanism.
GO:0042802 identical protein binding
IEA
GO_REF:0000118 		MARK AS OVER ANNOTATED
Summary: Self-association annotation reflecting the homodimeric quaternary structure typical of class-I aminotransferases (UniProt SUBUNIT: Homodimer). While the homodimer assignment is reasonable, "identical protein binding" is an uninformative interaction term that does not convey biological function and is a frequent TreeGrafter over-propagation.
Reason: Homodimerization is a structural property rather than a distinct molecular function; the term adds little and is propagated electronically without direct evidence for this protein. Per curation guidance, generic "protein binding"-type terms are discouraged.

Core Functions

PLP-dependent aromatic-amino-acid aminotransferase catalyzing reversible transamination between aromatic amino acids (L-tyrosine, L-phenylalanine) and their 2-oxoacids using 2-oxoglutarate/L-glutamate as the amino acceptor/donor pair, functioning in aromatic amino acid biosynthesis and catabolism.

Molecular Function:
L-tyrosine:2-oxoglutarate transaminase activity
Directly Involved In:
aromatic amino acid metabolic process
Supporting Evidence:

References

GO_REF:0000002
Gene Ontology annotation through association of InterPro records with GO terms
GO_REF:0000118
TreeGrafter-generated GO annotations
GO_REF:0000120
Combined Automated Annotation using Multiple IEA Methods
PMID:23685963
tyrB-2 and phhC genes of Pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level
  • P. putida aromatic aminotransferase isozymes preferentially transaminate aromatic amino acids and aromatic 2-oxoacids (best substrates L-phenylalanine and phenylpyruvate), using PLP cofactor and 2-oxoglutarate as amino acceptor.
PMID:20050871
Identification and characterization of the PhhR regulon in Pseudomonas putida
  • Genetic study of aromatic amino acid catabolism in P. putida KT2440; tyrB-1 (PP_1972) and tyrB-2 mutants show altered doubling times on tyrosine and phenylalanine as nitrogen sources, implicating tyrB-family aminotransferases in aromatic amino acid utilization.
PMID:38323821
Machine learning analysis of RB-TnSeq fitness data predicts functional gene modules in Pseudomonas putida KT2440
  • Disruption of tyrB (PP_1972) did not inhibit growth on L-phenylalanine or L-tyrosine as sole nitrogen sources, whereas disruption of AmaC (PP_3590) abolished growth; the authors propose PP_3590 as the dominant L-tyrosine aminotransferase, indicating PP_1972 is functionally redundant under those conditions.
PMID:35285712
Nitrogen metabolism in Pseudomonas putida: functional analysis using random barcode transposon sequencing
  • RB-TnSeq fitness data show only weak single-gene fitness effects for PP_1972 on aromatic nitrogen sources, and a PP_3590/PP_1972 double knockout did not cause phenylalanine auxotrophy, consistent with redundancy among aromatic aminotransferases.

Suggested Questions for Experts

Q: What is the in vitro substrate range and kinetic preference of purified PP_1972 (Tyr vs Phe vs Trp; 2-oxoglutarate vs pyruvate as acceptor), given that direct enzymology exists for the paralogs but not for PP_1972 itself?

Q: What is the division of labor among the P. putida KT2440 aromatic aminotransferase isozymes (PP_1972/tyrB-1, PP_3590/AmaC, tyrB-2/phhC) in aromatic amino acid biosynthesis versus catabolism, and under what conditions is PP_1972 non-redundant?

Suggested Experiments

Experiment: Express and purify recombinant PP_1972 and determine kinetic constants (Km/kcat) against a panel of amino donors (Tyr, Phe, Trp, Asp) and 2-oxoacid acceptors to define substrate specificity directly.

Experiment: Construct single and combinatorial in-frame deletions of PP_1972, PP_3590, and tyrB-2/phhC and assay growth on aromatic amino acids as sole nitrogen and carbon sources to resolve the redundancy network and any condition-specific, non-redundant role of PP_1972.

Deep Research

Falcon

(tyrB-deep-research-falcon.md)
Research report: **tyrB (PP_1972; UniProt Q88LG1)** in *Pseudomonas putida* KT2440 Falcon Edison Scientific Literature 19 citations 3 artifacts 2026-06-11T22:05:14.247139
Edison artifact artifact-00 (text/markdown)
## Context ID: pqac-00000009 Table 3 on page 5 reports the doubling times (growth phenotypes) of P. putida KT2440 and its isogenic mutants, including the `tyrB- (image/png)
## Context ID: pqac-00000026 The substrate specificity for ArAT I (tyrB-2) and ArAT II (phhC), including activities with aromatic amino acids and various 2-oxoa (image/png)

The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.

You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.

We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.

We are interested in where in or outside the cell the gene product carries out its function.

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Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.

Research report: tyrB (PP_1972; UniProt Q88LG1) in Pseudomonas putida KT2440

0) Target verification (gene/protein identity)

The requested UniProt entry Q88LG1 corresponds to Pseudomonas putida KT2440 gene tyrB with locus tag PP_1972. In KT2440-focused literature analyzing aromatic amino acid catabolism, PP1972 is explicitly referred to as tyrB-1, while a paralogous aminotransferase gene (tyrB-2) is associated with PP_3590 (also discussed as amaC in later work), confirming that the report targets the correct gene/protein in the correct organism/strain (ATCC 47054/KT2440) and not a different “tyrB” from another organism. (herrera2010identificationandcharacterization pages 1-2, borchert2024machinelearninganalysis pages 7-11)

1) Key concepts and definitions (current understanding)

1.1 What “TyrB” typically denotes in bacteria

In many bacteria, TyrB denotes a PLP-dependent aminotransferase that catalyzes reversible transamination reactions, transferring an amino group between an amino acid and an α-keto acid. In Pseudomonas, enzymes annotated as TyrB-family aminotransferases have been studied primarily in the context of aromatic amino acid transformations, i.e., interconversion between aromatic amino acids (phenylalanine/tyrosine/tryptophan) and their corresponding aromatic 2-oxoacids. (szkop2013tyrb2andphhc pages 1-2)

1.2 General PLP-aminotransferase chemistry (mechanism-level view)

PLP-dependent transaminases operate through two half-reactions in which the cofactor cycles between pyridoxal-5′-phosphate (PLP) and pyridoxamine phosphate (PMP), with key intermediates (external aldimine, quinonoid, ketimine) formed during amino-group transfer. A frequent structural theme is an oligomeric enzyme (often a homodimer) with active sites formed by residues from both subunits. (menke2024proteinengineeringof pages 22-25, menke2024proteinengineeringof pages 25-28)

This mechanism-level understanding is important for functional annotation because it explains (i) why PLP is required, (ii) why α-keto acids such as 2-oxoglutarate or pyruvate commonly serve as amino acceptors, and (iii) why these enzymes can show substrate promiscuity that complicates gene-to-function assignment by annotation alone. (menke2024proteinengineeringof pages 22-25, menke2024proteinengineeringof pages 25-28)

2) Molecular function of PP_1972 (tyrB; Q88LG1)

2.1 Enzyme class and likely reaction

Direct purified-enzyme biochemistry for PP_1972/Q88LG1 was not found in the retrieved full texts; however, multiple KT2440 studies place PP_1972 among aromatic/tyrosine aminotransferase-like genes and test its role genetically. (herrera2010identificationandcharacterization pages 1-2, herrera2010identificationandcharacterization pages 4-5, borchert2024machinelearninganalysis pages 7-11)

The reaction class most consistent with the TyrB annotation in this KT2440 context is an aromatic amino acid transamination such as:

  • L-tyrosine + 2-oxoglutarate ⇌ 4-hydroxyphenylpyruvate + L-glutamate
  • L-phenylalanine + 2-oxoglutarate ⇌ phenylpyruvate + L-glutamate

Support for this reaction type comes from protein-level characterization of closely related P. putida aromatic aminotransferases (encoded by tyrB-2 and phhC), which preferentially catalyze transamination involving aromatic amino acids and aromatic 2-oxoacids, with PLP included as cofactor and 2-oxoglutarate used as amino acceptor in assays. (szkop2013tyrb2andphhc pages 1-2, szkop2013tyrb2andphhc media f8f3824d)

2.2 Substrate specificity (direct evidence from P. putida aromatic aminotransferases)

While the enzyme characterized biochemically in P. putida was not PP_1972, Table 2 from Szkop & Bielawski (2013) provides detailed substrate profiles for two P. putida aromatic aminotransferase isozymes, showing that they most efficiently catalyze reactions involving aromatic amino acids and aromatic 2-oxoacids, with L-phenylalanine and phenylpyruvate being the best substrates reported. Assays used 0.1 M phosphate buffer (pH 8.0), 3 mM 2-oxoglutarate, 10 µM PLP, and 35 °C—conditions consistent with fold-type I PLP aminotransferase enzymology. (szkop2013tyrb2andphhc pages 1-2, szkop2013tyrb2andphhc media f8f3824d)

These data support that TyrB-like enzymes in P. putida are plausibly aromatic aminotransferases rather than strictly tyrosine-specific enzymes, and that substrate promiscuity and isozyme redundancy should be expected in vivo. (szkop2013tyrb2andphhc pages 1-2, szkop2013tyrb2andphhc media f8f3824d)

3) Biological role and pathways in P. putida KT2440

3.1 Phenylalanine utilization routes and PhhR-controlled regulon

A central KT2440 pathway context is phenylalanine assimilation/catabolism regulated by PhhR, which induces the phhAB operon encoding a pterin-dependent phenylalanine hydroxylase system (PhhA catalytic hydroxylase; PhhB pterin cofactor regeneration). This provides a route for conversion of L-phenylalanine to L-tyrosine. (herrera2010identificationandcharacterization pages 1-2)

Herrera et al. (2010) place phenylalanine/tyrosine degradation into a larger catabolic funnel via p-hydroxyphenylpyruvate and homogentisate (hpd/hmg genes), which then feeds into central metabolism. (herrera2010identificationandcharacterization pages 4-5)

3.2 Genetic evidence for tyrB-family involvement in phenylalanine/tyrosine catabolism

In KT2440, mutants in TyrB-like genes exhibit measurable growth phenotypes on aromatic amino acids as nitrogen sources.

  • Wild-type KT2440 doubling time: ~8 h on phenylalanine; ~1.8 h on tyrosine.
  • tyrB-1 (PP_1972) mutant: phenylalanine growth near WT, but slower on tyrosine (~3.2 h).
  • tyrB-2 mutant: slower on phenylalanine (~12 h) and slower on tyrosine (~3.0 h).

This pattern supports that TyrB-family enzymes contribute to aromatic amino acid utilization, with a stronger phenotype for tyrB-2 in phenylalanine conditions in this dataset. (herrera2010identificationandcharacterization media d9fda959, herrera2010identificationandcharacterization pages 4-5)

3.3 Redundancy and reassignment of “primary” aromatic aminotransferase (major recent development)

A key development in 2024 is that machine-learning analysis of RB-TnSeq fitness compendia (ICA-derived “fModules”) was used to pinpoint genes involved in phenylalanine/tyrosine catabolism, followed by mutant validation.

Borchert et al. (2024, published March 2024; https://doi.org/10.1128/msystems.00942-23) report that disruption of tyrB (PP_1972) did not inhibit growth on L-phenylalanine or L-tyrosine as sole nitrogen sources, whereas disruption of amaC (PP_3590; sometimes called tyrB2) completely abrogated growth on these substrates. The authors therefore propose re-annotation of AmaC (PP_3590) as an L-tyrosine aminotransferase, implying that PP_1972 is not the primary enzyme for these growth phenotypes under the tested conditions and that previous “tyrB” annotation may overstate its physiological importance. (borchert2024machinelearninganalysis pages 7-11)

This reconciles earlier BarSeq-based observations that PP_1972 often shows weak fitness effects and that a PP_3590/PP_1972 double knockout did not cause phenylalanine auxotrophy, consistent with broader redundancy or alternative routes, while still allowing PP_1972 to contribute in specific environments or regulatory states. (schmidt2022nitrogenmetabolismin pages 8-10, schmidt2022nitrogenmetabolismin pages 10-12)

4) Cellular localization

No retrieved KT2440 primary source in this corpus provided a direct experimental localization (cytosol/periplasm) for PP_1972/Q88LG1.

For context on what localization evidence looks like for bacterial PLP-dependent aminotransferases, Ringel et al. (2017) show that a distinct periplasmic PLP-dependent transaminase (PtaA) can be demonstrated by subcellular fractionation and is a homodimer by SEC-MALS; however, this is a different enzyme in a different Pseudomonas species and should not be taken as evidence that PP_1972 is periplasmic. (ringel2017theperiplasmictransaminase pages 16-18)

Current best-supported statement from the retrieved KT2440 corpus: localization of PP_1972 remains unresolved here and should be taken from UniProt/InterPro experimental annotations if available, or validated experimentally.

5) Current applications and real-world implementations

5.1 Functional genomics for strain engineering and annotation (2024)

The 2024 RB‑TnSeq + ICA framework provides a practical route to re-annotate metabolic genes and identify engineering targets for P. putida as a chassis. In particular, the ability to distinguish PP_1972 (tyrB) from PP_3590 (AmaC) as the functionally dominant aminotransferase for phenylalanine/tyrosine utilization under defined conditions is directly actionable for (i) redirecting aromatic amino acid flux, and (ii) avoiding incorrect knockouts when designing production strains. (borchert2024machinelearninganalysis pages 7-11)

5.2 Engineering relevance via aromatic-stress tolerance and aromatic feedstocks

Borchert et al. (2024) also connect aminotransferase-linked modules to tolerance phenotypes during growth with high concentrations of hydroxycinnamates (e.g., ~60 mM in glucose + hydroxycinnamate tests; and higher concentrations as carbon sources in some conditions). These results highlight that aromatic amino acid and aromatic acid metabolism genes can have roles in stress tolerance, a key trait for industrial bioprocessing on lignin-derived aromatics. (borchert2024machinelearninganalysis pages 7-11)

5.3 Broader industrial deployment of PLP-transaminases (expert review)

Menke (2024) reviews industrial and engineering aspects of PLP-dependent amine transaminases (ATAs), emphasizing (i) their use in stereoselective synthesis of chiral amines, (ii) the importance of addressing unfavorable equilibria (e.g., via coproduct removal or sacrificial donors), and (iii) the rise of machine-learning-guided engineering. The review notes an industrial benchmark: (R)-ATA-catalyzed synthesis of (R)-sitagliptin with >99.95% optical purity, and also reports large engineering gains (e.g., up to 2000-fold improved catalytic activity in one redesign example), illustrating the real-world value of understanding transaminase substrate specificity and engineering it rationally. (menke2024proteinengineeringof pages 61-71)

Although this review is not specific to PP_1972, it provides authoritative context for why TyrB-family enzymes and related aminotransferases are frequently targeted in metabolic engineering and biocatalysis.

6) Summary of key quantitative evidence (recent and classic)

  1. Mutant growth phenotypes (KT2440; 2010): doubling times on phenylalanine and tyrosine as sole N sources show measurable aromatic-amino-acid-related phenotypes for tyrB mutants, with a stronger phenotype for tyrB-2 in phenylalanine conditions. (herrera2010identificationandcharacterization media d9fda959)
  2. Functional genomics/fitness and redundancy (2022): PP_1972 shows weak fitness effects in some aromatic N-source conditions and is genetically redundant with other aminotransferases in some contexts. (schmidt2022nitrogenmetabolismin pages 8-10, schmidt2022nitrogenmetabolismin pages 10-12)
  3. Reannotation-driven experimental validation (2024): PP_1972 knockout does not inhibit growth on L-Phe/L-Tyr as sole N sources, while PP_3590 (AmaC) knockout abolishes growth, supporting updated functional assignment of the dominant aromatic aminotransferase in those conditions. (borchert2024machinelearninganalysis pages 7-11)
  4. Biochemical assay conditions and substrate profiles (2013): P. putida aromatic aminotransferase isozymes show strongest activity for aromatic amino acids/2-oxoacids (e.g., L-Phe/phenylpyruvate) in PLP + 2-oxoglutarate-dependent assays. (szkop2013tyrb2andphhc media f8f3824d)

7) Practical functional-annotation conclusion for PP_1972 (tyrB; Q88LG1)

Most defensible functional annotation from the retrieved evidence:

  • Gene/protein: tyrB (PP_1972; UniProt Q88LG1) in P. putida KT2440.
  • Molecular function: PLP-dependent aminotransferase in the aromatic/tyrosine aminotransferase-like set of enzymes in Pseudomonas; likely catalyzes reversible transamination between aromatic amino acids (e.g., tyrosine/phenylalanine) and α-keto acids (commonly 2-oxoglutarate), but direct biochemical constants for PP_1972 are not established in the retrieved corpus. (herrera2010identificationandcharacterization pages 1-2, szkop2013tyrb2andphhc media f8f3824d)
  • Biological process/pathway: participates (directly or redundantly) in aromatic amino acid utilization/catabolism connected to phenylalanine → tyrosine conversion (PhhAB) and downstream degradation via p-hydroxyphenylpyruvate/homogentisate. Genetic data indicate PP_1972 contributes measurably to tyrosine growth in some conditions but is not the primary determinant of Phe/Tyr utilization in others. (herrera2010identificationandcharacterization pages 4-5, herrera2010identificationandcharacterization media d9fda959, borchert2024machinelearninganalysis pages 7-11)
  • Systems-level role: part of a redundant aminotransferase network in KT2440; recent (2024) work indicates that PP_3590 (AmaC) dominates growth phenotypes on Phe/Tyr as N sources, and PP_1972 disruption alone may not block those pathways under standard minimal-media conditions. (borchert2024machinelearninganalysis pages 7-11, schmidt2022nitrogenmetabolismin pages 10-12)
  • Cellular localization: not resolved from retrieved KT2440 evidence; should be verified experimentally or via curated UniProt localization annotations. (ringel2017theperiplasmictransaminase pages 16-18)

8) Evidence table (quick reference)

Aspect Evidence summary Key quantitative data
Target identity UniProt Q88LG1 corresponds to tyrB / PP_1972 in Pseudomonas putida KT2440; genome annotation in KT2440 literature lists PP1972 as tyrB-1, one of two tyrosine/aromatic aminotransferase-like genes in this strain (herrera2010identificationandcharacterization pages 1-2, herrera2010identificationandcharacterization pages 9-10) Locus tags/names reported as PP1972 / tyrB-1; paralog also noted as PP3590 / tyrB-2 (herrera2010identificationandcharacterization pages 1-2)
Predicted molecular function/class TyrB/PP_1972 is an aminotransferase in the PLP-dependent aromatic amino acid aminotransferase class; related P. putida aromatic aminotransferases preferentially transaminate aromatic amino acids with 2-oxoglutarate, with best substrates including L-phenylalanine and phenylpyruvate (szkop2013tyrb2andphhc pages 2-4, szkop2013tyrb2andphhc pages 1-2) Assays for related P. putida ArAT enzymes used 10 µM PLP and 3 mM 2-oxoglutarate; activity measured as release of 1 µmol IPyA min⁻¹ in L-tryptophan:2-oxoglutarate assays (szkop2013tyrb2andphhc pages 2-4)
Pathway role in aromatic amino acid metabolism In KT2440, phenylalanine can be degraded by the phenylalanine hydroxylase pathway (PhhAB → tyrosine → p-hydroxyphenylpyruvate → homogentisate), and KT2440 carries two TyrB-like aminotransferase genes. Mutant phenotypes support TyrB-family participation in phenylalanine/tyrosine catabolism, especially downstream aromatic transamination steps (herrera2010identificationandcharacterization pages 4-5, herrera2010identificationandcharacterization pages 1-2) Wild type doubling times on sole N source: phenylalanine ~8 h, tyrosine ~1.8 h; tyrB-1 mutant: phenylalanine ~WT, tyrosine ~3.2 h; tyrB-2 mutant: phenylalanine ~12 h, tyrosine ~3.0 h (herrera2010identificationandcharacterization pages 4-5, herrera2010identificationandcharacterization media d9fda959)
Evidence for redundancy Recent RB-TnSeq and prior knockout work indicate functional redundancy among aromatic aminotransferases in P. putida KT2440: PP_1972 has only weak single-gene phenotypes in some aromatic N-source conditions, and even combined loss with PP_3590 did not cause phenylalanine auxotrophy (schmidt2022nitrogenmetabolismin pages 8-10, schmidt2022nitrogenmetabolismin pages 10-12) BarSeq fitness effects for PP_1972 were small: phenylalanine -0.35 and pipecolate -0.15 in one report; another excerpt summarizes similarly weak effects and cites no phenylalanine auxotrophy in the PP_3590 PP_1972 double knockout (schmidt2022nitrogenmetabolismin pages 8-10, schmidt2022nitrogenmetabolismin pages 10-12)
Strength/limits of direct evidence for Q88LG1 Evidence for PP_1972/Q88LG1 specifically is mainly genetic/fitness-based in KT2440; direct biochemical characterization in P. putida has more clearly identified other aromatic aminotransferase isozymes (tyrB-2/phhC) than PP_1972 itself, so annotation of Q88LG1 is supported by homology plus mutant evidence rather than purified-enzyme kinetics (szkop2013tyrb2andphhc pages 1-2, schmidt2022nitrogenmetabolismin pages 10-12) No direct purified-enzyme kinetic constants for PP_1972/Q88LG1 were extracted from the cited KT2440 sources; strongest KT2440-specific quantitative data are mutant doubling times and RB-TnSeq fitness values (schmidt2022nitrogenmetabolismin pages 10-12, herrera2010identificationandcharacterization media d9fda959)

Table: This table summarizes the strongest available evidence for functional annotation of Pseudomonas putida KT2440 tyrB (PP_1972; UniProt Q88LG1), including its identity, predicted aminotransferase role, pathway context, redundancy, and the key quantitative phenotypes available from mutant and fitness studies.

9) Source list (with URLs and publication dates where available)

  • Herrera MC, Duque E, Rodríguez‑Herva JJ, Fernández‑Escamilla AM, Ramos JL. Identification and characterization of the PhhR regulon in Pseudomonas putida. Environmental Microbiology (Jun 2010). https://doi.org/10.1111/j.1462-2920.2009.02124.x (herrera2010identificationandcharacterization pages 1-2, herrera2010identificationandcharacterization media d9fda959)
  • Szkop M, Bielawski W. tyrB-2 and phhC genes of Pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level. Amino Acids (May 2013). https://doi.org/10.1007/s00726-013-1508-y (szkop2013tyrb2andphhc pages 1-2, szkop2013tyrb2andphhc media f8f3824d)
  • Schmidt M et al. Nitrogen metabolism in Pseudomonas putida: functional analysis using random barcode transposon sequencing. Applied and Environmental Microbiology (Apr 2022). https://doi.org/10.1128/aem.02430-21 (schmidt2022nitrogenmetabolismin pages 8-10, schmidt2022nitrogenmetabolismin pages 10-12)
  • Borchert AJ et al. Machine learning analysis of RB‑TnSeq fitness data predicts functional gene modules in Pseudomonas putida KT2440. mSystems (Mar 2024). https://doi.org/10.1128/msystems.00942-23 (borchert2024machinelearninganalysis pages 7-11)
  • Menke M. Protein Engineering of Amine Transaminases and Methyltransferases using Machine Learning and High-Throughput Screening Tools. (2024; journal not captured in retrieved metadata). (menke2024proteinengineeringof pages 61-71, menke2024proteinengineeringof pages 25-28)
  • Direct PP_1972 enzymology: The retrieved corpus lacks purified-enzyme kinetics (Km/kcat) and direct substrate panels for PP_1972/Q88LG1 specifically; current enzymatic claims for PP_1972 are therefore best stated as inference from gene context + mutant/fitness phenotypes and from biochemical characterization of close isozymes in P. putida. (borchert2024machinelearninganalysis pages 7-11, szkop2013tyrb2andphhc media f8f3824d)
  • Localization: No KT2440 localization evidence for PP_1972 was found here. If localization is critical (e.g., for pathway compartmentalization or cofactor supply), it should be determined experimentally (cell fractionation, fluorescence tagging) or taken from curated UniProt/InterPro annotations.

References

  1. (herrera2010identificationandcharacterization pages 1-2): M. Carmen Herrera, Estrella Duque, José J. Rodríguez‐Herva, Ana M. Fernández‐Escamilla, and Juan L. Ramos. Identification and characterization of the phhr regulon in pseudomonas putida. Environmental microbiology, 12 6:1427-38, Jun 2010. URL: https://doi.org/10.1111/j.1462-2920.2009.02124.x, doi:10.1111/j.1462-2920.2009.02124.x. This article has 43 citations and is from a domain leading peer-reviewed journal.

  2. (borchert2024machinelearninganalysis pages 7-11): Andrew J. Borchert, Alissa C. Bleem, Hyun Gyu Lim, Kevin Rychel, Keven D. Dooley, Zoe A. Kellermyer, Tracy L. Hodges, Bernhard O. Palsson, and Gregg T. Beckham. Machine learning analysis of rb-tnseq fitness data predicts functional gene modules in pseudomonas putida kt2440. Mar 2024. URL: https://doi.org/10.1128/msystems.00942-23, doi:10.1128/msystems.00942-23. This article has 13 citations and is from a peer-reviewed journal.

  3. (szkop2013tyrb2andphhc pages 1-2): Michał Szkop and Wiesław Bielawski. Tyrb-2 and phhc genes of pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level. Amino Acids, 45:351-358, May 2013. URL: https://doi.org/10.1007/s00726-013-1508-y, doi:10.1007/s00726-013-1508-y. This article has 5 citations and is from a peer-reviewed journal.

  4. (menke2024proteinengineeringof pages 22-25): M Menke. Protein engineering of amine transaminases and methyltransferases using machine learning and high-throughput screening tools. Unknown journal, 2024.

  5. (menke2024proteinengineeringof pages 25-28): M Menke. Protein engineering of amine transaminases and methyltransferases using machine learning and high-throughput screening tools. Unknown journal, 2024.

  6. (herrera2010identificationandcharacterization pages 4-5): M. Carmen Herrera, Estrella Duque, José J. Rodríguez‐Herva, Ana M. Fernández‐Escamilla, and Juan L. Ramos. Identification and characterization of the phhr regulon in pseudomonas putida. Environmental microbiology, 12 6:1427-38, Jun 2010. URL: https://doi.org/10.1111/j.1462-2920.2009.02124.x, doi:10.1111/j.1462-2920.2009.02124.x. This article has 43 citations and is from a domain leading peer-reviewed journal.

  7. (szkop2013tyrb2andphhc media f8f3824d): Michał Szkop and Wiesław Bielawski. Tyrb-2 and phhc genes of pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level. Amino Acids, 45:351-358, May 2013. URL: https://doi.org/10.1007/s00726-013-1508-y, doi:10.1007/s00726-013-1508-y. This article has 5 citations and is from a peer-reviewed journal.

  8. (herrera2010identificationandcharacterization media d9fda959): M. Carmen Herrera, Estrella Duque, José J. Rodríguez‐Herva, Ana M. Fernández‐Escamilla, and Juan L. Ramos. Identification and characterization of the phhr regulon in pseudomonas putida. Environmental microbiology, 12 6:1427-38, Jun 2010. URL: https://doi.org/10.1111/j.1462-2920.2009.02124.x, doi:10.1111/j.1462-2920.2009.02124.x. This article has 43 citations and is from a domain leading peer-reviewed journal.

  9. (schmidt2022nitrogenmetabolismin pages 8-10): Matthias Schmidt, Allison N. Pearson, Matthew R. Incha, Mitchell G. Thompson, Edward E. K. Baidoo, Ramu Kakumanu, Aindrila Mukhopadhyay, Patrick M. Shih, Adam M. Deutschbauer, Lars M. Blank, and Jay D. Keasling. Nitrogen metabolism in pseudomonas putida: functional analysis using random barcode transposon sequencing. Applied and Environmental Microbiology, Apr 2022. URL: https://doi.org/10.1128/aem.02430-21, doi:10.1128/aem.02430-21. This article has 36 citations and is from a peer-reviewed journal.

  10. (schmidt2022nitrogenmetabolismin pages 10-12): Matthias Schmidt, Allison N. Pearson, Matthew R. Incha, Mitchell G. Thompson, Edward E. K. Baidoo, Ramu Kakumanu, Aindrila Mukhopadhyay, Patrick M. Shih, Adam M. Deutschbauer, Lars M. Blank, and Jay D. Keasling. Nitrogen metabolism in pseudomonas putida: functional analysis using random barcode transposon sequencing. Applied and Environmental Microbiology, Apr 2022. URL: https://doi.org/10.1128/aem.02430-21, doi:10.1128/aem.02430-21. This article has 36 citations and is from a peer-reviewed journal.

  11. (ringel2017theperiplasmictransaminase pages 16-18): Michael T. Ringel, Gerald Dräger, and Thomas Brüser. The periplasmic transaminase ptaa of pseudomonas fluorescens converts the glutamic acid residue at the pyoverdine fluorophore to α-ketoglutaric acid. Journal of Biological Chemistry, 292:18660-18671, Nov 2017. URL: https://doi.org/10.1074/jbc.m117.812545, doi:10.1074/jbc.m117.812545. This article has 17 citations and is from a domain leading peer-reviewed journal.

  12. (menke2024proteinengineeringof pages 61-71): M Menke. Protein engineering of amine transaminases and methyltransferases using machine learning and high-throughput screening tools. Unknown journal, 2024.

  13. (herrera2010identificationandcharacterization pages 9-10): M. Carmen Herrera, Estrella Duque, José J. Rodríguez‐Herva, Ana M. Fernández‐Escamilla, and Juan L. Ramos. Identification and characterization of the phhr regulon in pseudomonas putida. Environmental microbiology, 12 6:1427-38, Jun 2010. URL: https://doi.org/10.1111/j.1462-2920.2009.02124.x, doi:10.1111/j.1462-2920.2009.02124.x. This article has 43 citations and is from a domain leading peer-reviewed journal.

  14. (szkop2013tyrb2andphhc pages 2-4): Michał Szkop and Wiesław Bielawski. Tyrb-2 and phhc genes of pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level. Amino Acids, 45:351-358, May 2013. URL: https://doi.org/10.1007/s00726-013-1508-y, doi:10.1007/s00726-013-1508-y. This article has 5 citations and is from a peer-reviewed journal.

Artifacts

Citations

  1. herrera2010identificationandcharacterization pages 1-2
  2. herrera2010identificationandcharacterization pages 4-5
  3. borchert2024machinelearninganalysis pages 7-11
  4. ringel2017theperiplasmictransaminase pages 16-18
  5. menke2024proteinengineeringof pages 61-71
  6. menke2024proteinengineeringof pages 22-25
  7. menke2024proteinengineeringof pages 25-28
  8. schmidt2022nitrogenmetabolismin pages 8-10
  9. schmidt2022nitrogenmetabolismin pages 10-12
  10. herrera2010identificationandcharacterization pages 9-10
  11. https://doi.org/10.1128/msystems.00942-23
  12. https://doi.org/10.1111/j.1462-2920.2009.02124.x
  13. https://doi.org/10.1007/s00726-013-1508-y
  14. https://doi.org/10.1128/aem.02430-21
  15. https://doi.org/10.1111/j.1462-2920.2009.02124.x,
  16. https://doi.org/10.1128/msystems.00942-23,
  17. https://doi.org/10.1007/s00726-013-1508-y,
  18. https://doi.org/10.1128/aem.02430-21,
  19. https://doi.org/10.1074/jbc.m117.812545,

📄 View Raw YAML

 
id: Q88LG1
gene_symbol: tyrB
product_type: PROTEIN
status: DRAFT
taxon:
  id: NCBITaxon:160488
  label: Pseudomonas putida (strain ATCC 47054 / DSM 6125 / CFBP 8728 / NCIMB 11950 / KT2440)
description: >-
  tyrB (PP_1972; also referred to as tyrB-1) is a cytoplasmic, pyridoxal
  5'-phosphate (PLP)-dependent aminotransferase of the class-I (fold-type I)
  aspartate aminotransferase superfamily. It catalyzes reversible transamination
  in which an amino group is transferred from an amino acid donor to a 2-oxoacid
  acceptor (commonly 2-oxoglutarate, yielding L-glutamate). It is annotated as an
  aromatic-amino-acid aminotransferase, interconverting aromatic amino acids
  (L-tyrosine, L-phenylalanine) and their cognate aromatic 2-oxoacids
  (4-hydroxyphenylpyruvate, phenylpyruvate), and contributes to aromatic amino
  acid biosynthesis and catabolism. Like many class-I PLP aminotransferases it is
  a homodimer with active sites formed at the subunit interface. In P. putida
  KT2440 it is one of several aminotransferase isozymes acting on aromatic amino
  acids; genetic studies show that loss of tyrB alone causes only mild
  aromatic-amino-acid utilization phenotypes because of redundancy with paralogous
  aminotransferases (notably PP_3590/AmaC and tyrB-2/phhC), so its physiological
  role overlaps with those enzymes.
existing_annotations:
- term:
    id: GO:0003824
    label: catalytic activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: enables
  review:
    summary: >-
      Generic root-level molecular function term. tyrB is an enzyme, so this is
      not wrong, but it is uninformatively general and is fully subsumed by the
      more specific transaminase/aminotransferase activity terms.
    action: MARK_AS_OVER_ANNOTATED
    reason: >-
      "catalytic activity" is the MF root and conveys no specific information.
      The more precise terms GO:0008483 (transaminase activity) and GO:0004838
      (L-tyrosine:2-oxoglutarate transaminase activity) capture the actual
      function.
- term:
    id: GO:0004838
    label: L-tyrosine:2-oxoglutarate transaminase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000118
  qualifier: enables
  review:
    summary: >-
      Specific aromatic (tyrosine) aminotransferase activity assigned by
      TreeGrafter from the PTHR11879:SF37 "aromatic-amino-acid aminotransferase"
      subfamily. This is consistent with the protein family, the COG1448
      assignment, and with biochemical/genetic characterization of P. putida
      aromatic aminotransferases. This is the best representation of the gene's
      core molecular function.
    action: ACCEPT
    reason: >-
      Domain/family evidence (class-I PLP aminotransferase, PANTHER ArAT
      subfamily SF37) plus genetic evidence in KT2440 (tyrB mutants show a
      tyrosine-utilization phenotype; PMID:20050871) support tyrosine
      aminotransferase activity. The enzyme is likely promiscuous across aromatic
      amino acids (also acting on phenylalanine), but this term well captures the
      central characterized activity.
- term:
    id: GO:0005829
    label: cytosol
  evidence_type: IEA
  original_reference_id: GO_REF:0000118
  qualifier: located_in
  review:
    summary: >-
      Cytosolic localization predicted by TreeGrafter. Soluble class-I PLP
      aminotransferases acting in amino acid metabolism are cytoplasmic enzymes;
      the sequence has no signal peptide or transmembrane region. Consistent with
      the expected localization.
    action: ACCEPT
    reason: >-
      Aromatic aminotransferases in this family are soluble cytoplasmic enzymes.
      No experimental KT2440 localization data exist, but the prediction is
      biologically appropriate and there is no evidence for periplasmic/membrane
      localization.
- term:
    id: GO:0006520
    label: amino acid metabolic process
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: involved_in
  review:
    summary: >-
      Broad biological process term. tyrB participates in (aromatic) amino acid
      metabolism, so this is correct but general. A more specific process such as
      aromatic amino acid family metabolism / phenylalanine or tyrosine
      biosynthesis or catabolism would be more informative.
    action: MODIFY
    reason: >-
      The annotation is correct in essence but too high-level. tyrB acts
      specifically on aromatic amino acids (Tyr/Phe), so the more specific
      "aromatic amino acid family metabolic process" better reflects the
      characterized role while remaining defensible from family + genetic
      evidence. (Chorismate metabolic process is not appropriate: tyrB acts
      downstream of chorismate on the aromatic amino acids/2-oxoacids, not on
      chorismate itself.)
    proposed_replacement_terms:
    - id: GO:0009072
      label: aromatic amino acid family metabolic process
- term:
    id: GO:0008483
    label: transaminase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: enables
  review:
    summary: >-
      General transaminase (aminotransferase) activity. Correct and well
      supported by the class-I PLP-dependent aminotransferase family assignment,
      but less specific than GO:0004838. Useful as a parent term.
    action: KEEP_AS_NON_CORE
    reason: >-
      Accurately describes the enzymatic class but is a parent of the more
      specific aromatic aminotransferase term that represents the core function.
      Retain as supporting/non-core rather than as the primary MF.
- term:
    id: GO:0030170
    label: pyridoxal phosphate binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: enables
  review:
    summary: >-
      PLP cofactor binding. tyrB is a PLP-dependent enzyme (UniProt COFACTOR:
      pyridoxal 5'-phosphate; conserved PROSITE PS00105 class-I aminotransferase
      PLP-binding motif). This is a well-supported and informative molecular
      function annotation.
    action: ACCEPT
    reason: >-
      Strong family/motif evidence (IPR004838/IPR004839, PROSITE AA_TRANSFER_
      CLASS_1, UniProt cofactor annotation) for PLP binding, which is essential
      for the transamination mechanism.
- term:
    id: GO:0042802
    label: identical protein binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000118
  qualifier: enables
  review:
    summary: >-
      Self-association annotation reflecting the homodimeric quaternary structure
      typical of class-I aminotransferases (UniProt SUBUNIT: Homodimer). While
      the homodimer assignment is reasonable, "identical protein binding" is an
      uninformative interaction term that does not convey biological function and
      is a frequent TreeGrafter over-propagation.
    action: MARK_AS_OVER_ANNOTATED
    reason: >-
      Homodimerization is a structural property rather than a distinct molecular
      function; the term adds little and is propagated electronically without
      direct evidence for this protein. Per curation guidance, generic
      "protein binding"-type terms are discouraged.
core_functions:
- description: >-
    PLP-dependent aromatic-amino-acid aminotransferase catalyzing reversible
    transamination between aromatic amino acids (L-tyrosine, L-phenylalanine) and
    their 2-oxoacids using 2-oxoglutarate/L-glutamate as the amino acceptor/donor
    pair, functioning in aromatic amino acid biosynthesis and catabolism.
  molecular_function:
    id: GO:0004838
    label: L-tyrosine:2-oxoglutarate transaminase activity
  supported_by:
  - reference_id: PMID:23685963
  - reference_id: PMID:20050871
  directly_involved_in:
  - id: GO:0009072
    label: aromatic amino acid metabolic process
proposed_new_terms: []
suggested_questions:
- question: >-
    What is the in vitro substrate range and kinetic preference of purified
    PP_1972 (Tyr vs Phe vs Trp; 2-oxoglutarate vs pyruvate as acceptor), given
    that direct enzymology exists for the paralogs but not for PP_1972 itself?
- question: >-
    What is the division of labor among the P. putida KT2440 aromatic
    aminotransferase isozymes (PP_1972/tyrB-1, PP_3590/AmaC, tyrB-2/phhC) in
    aromatic amino acid biosynthesis versus catabolism, and under what conditions
    is PP_1972 non-redundant?
suggested_experiments:
- description: >-
    Express and purify recombinant PP_1972 and determine kinetic constants
    (Km/kcat) against a panel of amino donors (Tyr, Phe, Trp, Asp) and 2-oxoacid
    acceptors to define substrate specificity directly.
- description: >-
    Construct single and combinatorial in-frame deletions of PP_1972, PP_3590,
    and tyrB-2/phhC and assay growth on aromatic amino acids as sole nitrogen and
    carbon sources to resolve the redundancy network and any condition-specific,
    non-redundant role of PP_1972.
references:
- id: GO_REF:0000002
  title: Gene Ontology annotation through association of InterPro records with GO terms
  findings: []
- id: GO_REF:0000118
  title: TreeGrafter-generated GO annotations
  findings: []
- id: GO_REF:0000120
  title: Combined Automated Annotation using Multiple IEA Methods
  findings: []
- id: PMID:23685963
  title: "tyrB-2 and phhC genes of Pseudomonas putida encode aromatic amino acid aminotransferase isozymes: evidence at the protein level"
  findings:
  - statement: >-
      P. putida aromatic aminotransferase isozymes preferentially transaminate
      aromatic amino acids and aromatic 2-oxoacids (best substrates L-phenylalanine
      and phenylpyruvate), using PLP cofactor and 2-oxoglutarate as amino acceptor.
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      PMID verified via PubMed (Szkop & Bielawski, Amino Acids 2013). Establishes
      aromatic aminotransferase activity for P. putida isozymes (paralogs of
      PP_1972), supporting the family-level molecular function assignment.
- id: PMID:20050871
  title: "Identification and characterization of the PhhR regulon in Pseudomonas putida"
  findings:
  - statement: >-
      Genetic study of aromatic amino acid catabolism in P. putida KT2440; tyrB-1
      (PP_1972) and tyrB-2 mutants show altered doubling times on tyrosine and
      phenylalanine as nitrogen sources, implicating tyrB-family aminotransferases
      in aromatic amino acid utilization.
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      PMID verified via PubMed (Herrera et al., Environ Microbiol 2010). Provides
      KT2440-specific genetic evidence linking PP_1972 to aromatic amino acid
      metabolism.
- id: PMID:38323821
  title: "Machine learning analysis of RB-TnSeq fitness data predicts functional gene modules in Pseudomonas putida KT2440"
  findings:
  - statement: >-
      Disruption of tyrB (PP_1972) did not inhibit growth on L-phenylalanine or
      L-tyrosine as sole nitrogen sources, whereas disruption of AmaC (PP_3590)
      abolished growth; the authors propose PP_3590 as the dominant L-tyrosine
      aminotransferase, indicating PP_1972 is functionally redundant under those
      conditions.
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      PMID verified via PubMed (Borchert et al., mSystems 2024). Important for
      interpreting the in-vivo, non-core/redundant role of PP_1972.
- id: PMID:35285712
  title: "Nitrogen metabolism in Pseudomonas putida: functional analysis using random barcode transposon sequencing"
  findings:
  - statement: >-
      RB-TnSeq fitness data show only weak single-gene fitness effects for
      PP_1972 on aromatic nitrogen sources, and a PP_3590/PP_1972 double knockout
      did not cause phenylalanine auxotrophy, consistent with redundancy among
      aromatic aminotransferases.
  reference_review:
    relevance: MEDIUM
    correctness: VERIFIED
    review_notes: >-
      PMID verified via PubMed (Schmidt et al., Appl Environ Microbiol 2022).
      Supports the redundancy interpretation; corroborating rather than primary.