lhpL (PA1255, Q9I489) notes

2026-10-28 review (PseudoCAP IBA-seed worklist, rank 86)

No deep-research provider keys were available; literature gathered by PubMed
search and from the cached publications. Sister genes in the same hydroxyproline
cluster are not yet reviewed.

Identity

• UniProt Q9I489 (Swiss-Prot, PE 1), 344 aa, RecName "Bifunctional
  trans-3-hydroxy-L-proline dehydratase/2-epimerase", gene lhpL /
  OrderedLocusNames PA1255 [file:PSEAE/lhpL/lhpL-uniprot.txt].
• Proline racemase family (Pfam PF05544 Pro_racemase, PIRSF029792,
  SFLDS00028 Proline_Racemase, NCBIfam NF047722 T3LHypDht, PANTHER
  PTHR33442:SF5). Catalytic Cys-Cys pair architecture of that family; UniProt
  places the proton acceptor at residue 90 by similarity to B9K4G4
  [file:PSEAE/lhpL/lhpL-uniprot.txt].
• Two distinct enzymes must be kept apart in PAO1:
• PA1268 (Q9I476) is the genuine 4-hydroxyproline 2-epimerase
  (4HypE/PaHyPRE, EC 5.1.1.8).
• PA1255 (Q9I489, this gene) acts on the 3-hydroxy isomer.
  PseudoCAP annotated GO:0047580 (4-hydroxyproline epimerase activity) to both
  from PMID:27145750 (QuickGO, reference PMID:27145750).

Activity

• Watanabe et al. 2016 assign the 3-hydroxy activity to this gene
  PMID:27929065. That
  paper's own subject is the aconitase-X-family dehydratase PaLhpI, which acts
  on cis-3-hydroxy-L-proline; PaLhpL is the companion enzyme for the trans
  isomer.
• UniProt records both reactions with experimental evidence
  (ECO:0000269|PubMed:27929065): dehydration of trans-3-hydroxy-L-proline to
  Delta(1)-pyrroline-2-carboxylate (EC 4.2.1.77, KM 23 mM, kcat 28.7 min-1) and
  2-epimerization of trans-3-hydroxy-L-proline to cis-3-hydroxy-D-proline
  (EC 5.1.1.-) [file:PSEAE/lhpL/lhpL-uniprot.txt].
• The substrates it does not accept are also curated experimentally:
  "No dehydratase activity with L-proline, trans-4-hydroxy-L-" proline,
  "cis-4-hydroxy-L-proline (c4LHyp), D-proline" and so on, and, decisively for
  the seed under review, "Displays neither t4LHyp epimerase nor proline
  racemase activity (PubMed:17849014)"
  [file:PSEAE/lhpL/lhpL-uniprot.txt]. Goytia et al. assayed P. aeruginosa
  proline-racemase-family proteins directly and found that the ones with
  substituted catalytic couples gave neither activity
  PMID:17849014, while the genuine hydroxyproline epimerase they
  characterised (PaHyPRE) came from a different gene
  PMID:17849014.
• Zhao et al. 2014 is the genome-neighbourhood study that predicted t3HypD
  activities in this family; Q9I489 is among the sequences it treats
  PMID:24980702,
  and it is the source of the UniProt INDUCTION statement that lhpL is
  up-regulated on trans-4- or trans-3-hydroxy-L-proline as sole carbon source.

What PMID:27145750 (the seed reference) shows

• Li & Lu characterised the AraC-family regulator LhpR and the two hydroxyproline
  loci. In their nomenclature the gene at PA1255 is lhpK
  PMID:27145750; Hyp/Pro racemase is a
  sequence-based name, not an assay.
• The genetic result that PseudoCAP used is an induction phenotype of a double
  mutant PMID:27145750. That shows the two genes
  together are needed for D-Hyp-dependent promoter induction; it does not show
  which reaction either protein catalyses, and it cannot distinguish the 4- from
  the 3-hydroxy isomer. (Only the abstract was read; no PMC copy was found on
  2026-10-29, so other data in the full text are unchecked.)
• The term error itself does not depend on that paper: the full text of
  PMID:17849014 lists Q9I489 as the Ser-Cys (SC) variant in Table 1 and shows
  in Figure 1 that it has neither racemase nor hydroxyproline epimerase
  activity PMID:17849014. The two lhpL
  defects are therefore CONFIRMED (2026-10-29 audit).
• So the seed annotation is a phenotype-to-molecular-function step onto a term
  (GO:0047580: trans-4-hydroxy-L-proline = cis-4-hydroxy-D-proline) whose
  reaction has been directly tested on this protein and found absent.

PAINT node (lhpL-bioinformatics/RESULTS.md)

PTN002433936 is an internal DUPLICATION node of PTHR33442 (family root
PTN002530875, 48 leaves): 9 leaves in 7 organisms, sole-seeded by
UniProtKB:Q9I489 (PAINT snapshot 20180110). Leaves are proline-racemase-family
proteins of Korarchaeum, Chloroflexus, three Bacillus cereus paralogues,
Streptomyces coelicolor, Clostridium botulinum PrdF and Nematostella,
plus lhpL; one of the B. cereus leaves is itself named
"Trans-3-hydroxy-L-proline dehydratase". GOA currently holds 35 GO:0047580 rows
with this node in with/from (9 GO_Central reference leaves, 24 TreeGrafter,
2 UniProt), so the term the node carries is propagated well beyond the
reference proteome.

Annotation decisions

• GO:0047580 (IBA, IEA/ARBA, IMP): all three MODIFY to GO:0050346. The protein
  does have a hydroxyproline-acting epimerase/dehydratase activity, but on the
  3-hydroxy isomer; the 4-hydroxy reaction is experimentally excluded. The
  ARBA row's rule was almost certainly learnt from the same mis-assignment.
• GO:0050346 (EXP, IEA from RHEA/EC): ACCEPT, core.
• PSEUDOCAP WRONG_TERM defect on the IMP row, GO_CENTRAL WRONG_TERM defect
  on PTN002433936.
• The 2-epimerase half of the activity (trans-3-hydroxy-L-proline =
  cis-3-hydroxy-D-proline) has no GO term; recorded in proposed_new_terms.

PseudoCAP IBA-seed assessment