Gene: FSH3 / YOR280C, Saccharomyces cerevisiae (S288C), UniProt Q99369, SGD S000005806.
Name: "Family of Serine Hydrolases 3". 266 aa. UniProt EC=3.1.-.- (esterase, generic).
This is a functionally dark gene: a putative serine hydrolase of unknown physiological
substrate / reaction / in-vivo role.
KNOWN (defensible):
- Belongs to the α/β-hydrolase fold, AB-hydrolase-3 family; Pfam FSH1 (PF03959),
InterPro AB_hydrolase_fold (IPR029058) + FSH-like domain (IPR005645); PANTHER family
PTHR48070 subfamily SF6 "ESTERASE OVCA2"; SCOP/SUPFAM alpha/beta-Hydrolases (SSF53474).
- Has an intact, canonical serine-hydrolase catalytic triad (Ser-Asp-His) with the
nucleophile in a classic GXSXG motif (see domain analysis below).
- Was directly detected as an active serine hydrolase by activity-based protein profiling
(chemical + computational proteomics) in yeast — i.e. the catalytic serine is chemically
reactive in vivo PMID:14645503.
- Transcriptionally induced as a target of the Crt1/RFX1 repressor (DNA-damage-checkpoint
effector) PMID:15494396, and induced by H2O2/oxidative stress PMID:30776074.
- Localizes to the peroxisome (IDA, SGD) PMID:36164978; is one of a set of newly identified
peroxisomal proteins of unknown molecular function.
- Overexpression phenotypes: overexpression causes an apoptotic/ROS phenotype dependent on
NUC1 PMID:30776074, and overexpression produces the strongest lipidome change (reduction
of phosphatidylcholine, PC) among a panel of dark peroxisomal proteins PMID:36164978.
NOT KNOWN (the gaps — primary deliverable):
- The physiological substrate and the specific reaction it catalyzes. UniProt states
"Serine hydrolase of unknown specificity" and assigns only EC 3.1.-.- (uncommitted esterase).
- The in-vivo biological role / pathway. Single deletion has no reported growth phenotype
under standard conditions, but a double-deletion (Δlpx1Δfsh3) shows reduced peroxisomal
β-oxidation activity PMID:36164978, implicating FSH3 (redundantly with Lpx1) in
peroxisomal fatty-acid β-oxidation. The specific molecular step/substrate remains unassigned.
- Functional redundancy with paralogs FSH1 and FSH2 (double-deletion data below suggest
partial redundancy / opposing roles but do not assign a molecular function).
- Whether the PC reduction on overexpression reflects a direct enzymatic activity on a
glycerophospholipid substrate, or an indirect consequence.
UniProt annotates three "Charge relay system" active-site residues (by similarity, ECO:0000250):
- ACT_SITE 117, 180, 209.
Mapping onto the 266-aa sequence (verified computationally, see below):
- Position 117 = Ser (nucleophile), embedded in G115-F116-S117-Q118-G119 = GFSQG, a
textbook GXSXG "nucleophile elbow" motif of α/β-hydrolases.
- Position 180 = Asp (acid of the triad).
- Position 209 = His (base of the triad).
So FSH3 carries a complete, canonical Ser-Asp-His serine-hydrolase catalytic triad on an
α/β-hydrolase scaffold, with the nucleophilic serine in the diagnostic GXSXG motif. The triad
is intact (this is NOT a pseudoenzyme). Consistent with this, activity-based profiling detected
FSH3 as a catalytically active serine hydrolase in vivo PMID:14645503. This makes a generic
serine-hydrolase molecular function (GO:0017171 serine hydrolase activity) domain-defensible;
a specific substrate/reaction remains a genuine gap.
Verification (residue identity + motif), reproducible:
seq = ("MSEKKKVLMLHGFVQSDKIFSAKTGGLRKNLKKLGYDLYYPCAPHSIDKKALFQSESEKG"
"RDAAKEFNTSATSDEVYGWFFRNPESFNSFQIDQKVFNYLRNYVLENGPFDGVIGFSQGA"
"GLGGYLVTDFNRILNLTDEQQPALKFFISFSGFKLEDQSYQKEYHRIIQVPSLHVRGELD"
"EVVAESRIMALYESWPDNKRTLLVHPGAHFVPNSKPFVSQVCNWIQGITSKEGQEHNAQP"
"EVDRKQFDKPQLEDDLLDMIDSLGKL")
assert seq[116] == "S" and seq[179] == "D" and seq[208] == "H"
import re; assert re.search(r"G.S.G", seq).start()+1 == 115 # GFSQG, Ser at 117
S. cerevisiae has three FSH paralogs, all named for this "Family of Serine Hydrolases":
- FSH1 = P38777 / YHR049W (243 aa). PANTHER subfamily SF9 (distinct from FSH3).
The crystal structure 1YCD is FSH1/YHR049W, "a member of the serine hydrolase family"
(PANTHER PTHR48070 representative structure).
- FSH2 = Q05015 / YMR222C (223 aa). PANTHER subfamily SF6 (same subfamily as FSH3).
- FSH3 = Q99369 / YOR280C (266 aa). PANTHER subfamily SF6 "ESTERASE OVCA2".
So FSH3 is in the OVCA2 subfamily (SF6) together with FSH2 and human OVCA2, and is more
distantly related to FSH1 (SF9). The human ortholog is OVCA2 (Q8WZ82), a candidate ovarian
tumor-suppressor esterase; OVCA2 is the source of the phylogenetic (IBA) inferences in GOA
(GOA IBA uses UniProtKB:Q8WZ82 and PANTHER:PTN000512658 as with/from). The S. pombe member of
this family is DYR-SCHPO / SPAC22A12.06c.
Provenance for the family framing:
- PMID:14645503
- PMID:30776074
CAUTION: The DYR-SCHPO/DHFR association in PMID:14645503 and the speculation in
PMID:15494396 that "FSH3p ... may be involved in folate metabolism ... by carrying serine
hydrolase activity required for the novel metabolic pathway involving dihydrofolate reductase
(DHFR) or by directly interacting with the DHFR enzyme" is speculation from sequence
similarity, not experimental evidence for a FSH3 DHFR/folate function. Do not annotate folate
metabolism from this.
Activity-based/computational proteomics identifying active serine hydrolases in yeast; defines
the Fsh family and detects FSH3 as an active serine hydrolase. This is the source cited by
UniProt for FUNCTION "Serine hydrolase of unknown specificity" (ECO:0000269|PubMed:14645503).
- Supports: serine hydrolase activity (catalytically active in vivo), family definition.
- Abstract-only in cache (full_text_available: false).
FSH3 is a confirmed transcriptional target of Crt1p (RFX1), the DNA-damage-checkpoint effector.
- PMID:15494396
- The DHFR/folate idea here is speculative (see CAUTION above).
- Source of UniProt INDUCTION "Regulated by RFX1/CRT1".
Systematic multi-level analysis of the peroxisome proteome. FSH3 is a newly identified
peroxisomal protein (basis of the SGD IDA C:peroxisome annotation).
- PMID:36164978
- Supports: C:peroxisome (IDA); a lipid-metabolism-linked phenotype on overexpression
(reduction of phosphatidylcholine). Note this is an overexpression lipidome effect, not
proof of a direct PC-hydrolase activity.
- IMPORTANT (deletion-based phenotype, from full text): a peroxisomal β-oxidation activity
assay of single and double deletions shows Δfsh3 contributes to β-oxidation:
PMID:36164978. Lpx1 is the peroxisomal lipase; the double mutant Δlpx1Δfsh3 (but not either
single mutant) has reduced β-oxidation. This is a concrete deletion (loss-of-function)
clue that FSH3 has a redundant/overlapping role in peroxisomal fatty-acid β-oxidation —
consistent with the peroxisomal localization and the PC-reduction lipidome result. It
refines the "no deletion phenotype" statement: single-deletion growth is silent, but
a biochemical double-mutant β-oxidation phenotype exists and points to a candidate
pathway (GO:0006635 fatty acid beta-oxidation; in yeast this is exclusively peroxisomal).
This is why single fsh/lpx1 deletions look silent (redundancy). The molecular role of
FSH3 in that pathway (which acyl-ester substrate it hydrolyzes) is still unassigned.
Screen for hydrolytic enzymes in yeast peroxisomes and lipid droplets using GXSXG-motif serine
hydrolases; identifies Ayr1p as a TG lipase and confirms Lpx1p. Provides context for the class
of GXSXG serine hydrolases acting in peroxisomal/lipid metabolism (not a FSH3-specific paper).
- PMID:24187129
- Relevance: contextual (LOW/MEDIUM) — supports plausibility of a peroxisomal lipid-related
serine-hydrolase role, does not assign FSH3 a substrate.