dadX

UniProt ID: Q9HTQ2
Organism: Pseudomonas aeruginosa PAO1
Review Status: COMPLETE
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Gene Description

DadX (PA5302) is the catabolic alanine racemase of Pseudomonas aeruginosa, a pyridoxal 5'-phosphate-dependent homodimeric enzyme that interconverts L- and D-alanine (Km about 1.4 mM for each enantiomer). It works with the D-amino acid dehydrogenase DadA in L-alanine utilisation, converting L-alanine to D-alanine that DadA then oxidises to pyruvate. It is one of two alanine racemases in PAO1; the second, Alr, is the constitutive biosynthetic enzyme, although DadX can also supply D-alanine when expressed in an E. coli D-alanine auxotroph.

Existing Annotations Review

GO Term Evidence Action Reason
GO:0003824 catalytic activity
IEA
GO_REF:0000002 		TOO GENERAL
Summary: InterPro2GO from the alanine racemase/decarboxylase C-terminal superfamily.
Reason: True, but entailed by the accepted GO:0008784 alanine racemase activity.
GO:0005829 cytosol
IBA
GO_REF:0000033 		KEEP AS NON CORE
Summary: IBA from PTN000775508, seeded by E. coli DadX (EcoCyc IDA). DadX is a soluble enzyme with no signal peptide or membrane segment.
Reason: Plausible location for a soluble PLP enzyme, inherited from the E. coli ortholog; not part of the core function.
GO:0008784 alanine racemase activity
IBA
GO_REF:0000033 		ACCEPT
Summary: IBA from PTN000775508 with eight seeds, including this gene's own IDA row (expected self-appearance) and IDA-supported E. coli Alr, E. coli DadX and M. tuberculosis Alr. Sound at the node.
Reason: Core activity, consistent with the direct assay on purified DadX.
Supporting Evidence:
PMID:10977898
Both gene products were purified to electrophoretic homogeneity, the enzymes were characterized biochemically
GO:0008784 alanine racemase activity
IDA
PMID:10977898
Characterization of the alanine racemases from Pseudomonas a... 		ACCEPT
Summary: PseudoCAP seed row for PTN000775508. Purified DadX racemises alanine with measured kinetics. Seed verdict SOUND.
Reason: Direct assay on the purified enzyme; the core molecular function.
Supporting Evidence:
PMID:10977898
Both gene products were purified to electrophoretic homogeneity, the enzymes were characterized biochemically
file:PSEAE/dadX/dadX-uniprot.txt
Isomerizes L-alanine to D-alanine which is then oxidized to
GO:0008784 alanine racemase activity
IEA
GO_REF:0000120 		ACCEPT
Summary: Combined IEA from the alanine racemase family signature and EC 5.1.1.1.
Reason: Consistent with the experimental row.
Supporting Evidence:
PMID:10977898
Both gene products were purified to electrophoretic homogeneity, the enzymes were characterized biochemically
GO:0030170 pyridoxal phosphate binding
IBA
GO_REF:0000033 		ACCEPT
Summary: IBA from PTN000775508, seeded by E. coli and M. tuberculosis Alr (IDA).
Reason: PLP is the catalytic cofactor; the DadX crystal structure shows PLP bound at Lys33 (UniProt, ECO:0000269 PubMed:14674749).
Supporting Evidence:
file:PSEAE/dadX/dadX-uniprot.txt
Name=pyridoxal 5'-phosphate; Xref=ChEBI:CHEBI:597326;
GO:0030170 pyridoxal phosphate binding
IEA
GO_REF:0000104 		ACCEPT
Summary: UniRule transfer of PLP binding.
Reason: Consistent with the structure-based cofactor assignment.
Supporting Evidence:
file:PSEAE/dadX/dadX-uniprot.txt
Name=pyridoxal 5'-phosphate; Xref=ChEBI:CHEBI:597326;
GO:0030632 D-alanine biosynthetic process
IBA
GO_REF:0000033 		KEEP AS NON CORE
Summary: IBA from the family-level node PTN001253738, seeded by the biosynthetic Alr enzymes of E. coli and M. tuberculosis.
Reason: DadX does form D-alanine and complements E. coli D-alanine auxotrophs, so the claim holds, but its physiological role is L-alanine catabolism with DadA; peptidoglycan D-alanine is mainly the job of Alr.
Supporting Evidence:
PMID:10977898
The dadX(PA) and alr(PA) genes have been sequenced, overexpressed, and their activity was demonstrated by complementing d-alanine auxotrophs of Escherichia coli.
GO:0030632 D-alanine biosynthetic process
IEA
GO_REF:0000120 		KEEP AS NON CORE
Summary: Combined IEA from the alanine racemase family.
Reason: Same claim as the IBA row; true but not the main role of DadX.
Supporting Evidence:
PMID:10977898
The dadX(PA) and alr(PA) genes have been sequenced, overexpressed, and their activity was demonstrated by complementing d-alanine auxotrophs of Escherichia coli.
GO:0042853 L-alanine catabolic process
IC
file:PSEAE/dadX/dadX-uniprot.txt 		NEW
Summary: Proposed from the curated UniProt function statement (ECO:0000269, PubMed:10977898): DadX converts L-alanine to D-alanine for oxidation to pyruvate by DadA.
Reason: DadX performs the racemisation step of the DadX-DadA L-alanine utilisation route, so it does part of the work of the process. Not yet annotated in GOA.
Supporting Evidence:
file:PSEAE/dadX/dadX-uniprot.txt
Isomerizes L-alanine to D-alanine which is then oxidized to

Core Functions

PLP-dependent alanine racemase that converts L-alanine to D-alanine for oxidation by DadA in L-alanine catabolism.

Molecular Function:
alanine racemase activity
Directly Involved In:
L-alanine catabolic process
Supporting Evidence:
  • PMID:10977898
    Both gene products were purified to electrophoretic homogeneity, the enzymes were characterized biochemically
  • file:PSEAE/dadX/dadX-uniprot.txt
    Isomerizes L-alanine to D-alanine which is then oxidized to

References

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Suggested Questions for Experts

Q: Does DadX contribute measurably to peptidoglycan D-alanine in P. aeruginosa when alr is deleted, or is it confined to alanine catabolism?

Suggested Experiments

Experiment: Construct alr, dadX and alr dadX deletions in PAO1 and test D-alanine auxotrophy and growth on L-alanine as sole carbon source.

📚 Additional Documentation

Notes

(dadX-notes.md)

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