Gene: ACAA2 / 3-ketoacyl-CoA thiolase, mitochondrial / acetyl-CoA acyltransferase 2 / T1
UniProt: P42765 (THIM_HUMAN), 397 aa, EC 2.3.1.16
HGNC:83; GeneID 10449; chromosome 18.
ACAA2 catalyzes the fourth and last step of each round of the mitochondrial fatty acid
beta-oxidation spiral: the thiolytic (CoA-dependent) cleavage of a 3-ketoacyl-CoA
(3-oxoacyl-CoA) into acetyl-CoA and a fatty acyl-CoA shortened by two carbon atoms.
From UniProt FUNCTION (P42765):
- [UniProt:P42765 "this is one of the enzymes that catalyzes the last step of the mitochondrial beta-oxidation pathway, an aerobic process breaking down fatty acids into acetyl-CoA"]
- [UniProt:P42765 "Using free coenzyme A/CoA, catalyzes the thiolytic cleavage of medium- to long-chain unbranched 3-oxoacyl-CoAs into acetyl-CoA and a fatty acyl-CoA shortened by two carbon atoms"]
- Reverse/biosynthetic direction: [UniProt:P42765 "Also catalyzes the condensation of two acetyl-CoA molecules into acetoacetyl-CoA and could be involved in the production of ketone bodies"]
- Side hydrolase activity: [UniProt:P42765 "Also displays hydrolase activity on various fatty acyl-CoAs (PubMed:25478839). Thereby, could be responsible for the production of acetate in a side reaction to beta-oxidation"]
- Pathway: [UniProt:P42765 "PATHWAY: Lipid metabolism; fatty acid beta-oxidation."]
- Subunit: [UniProt:P42765 "Homotetramer (PubMed:25478839). Interacts with BNIP3."]
- Location: [UniProt:P42765 "SUBCELLULAR LOCATION: Mitochondrion"]; Reactome places it in mitochondrial matrix (soluble matrix enzyme, distinct from the membrane-bound MTP).
- Family: [UniProt:P42765 "Belongs to the thiolase-like superfamily. Thiolase family."]
EC numbers assigned by UniProt: EC 2.3.1.16 (3-ketoacyl-CoA thiolase / acetyl-CoA
C-acyltransferase), EC 2.3.1.9 (acetyl-CoA C-acetyltransferase, the acetoacetyl-CoA
synthesis/cleavage reaction), and EC 3.1.2.1 / 3.1.2.- (acyl-CoA hydrolase side activity).
The crystal structure (PDB 4C2J/4C2K, 2.0 Å) of human mitochondrial T1 (hT1 = ACAA2):
- PMID:25478839
- Active site resembles a biosynthetic tetrameric thiolase rather than the peroxisomal dimeric degradative thiolase: PMID:25478839
- Intrinsic thioesterase/hydrolase activity confirmed: PMID:25478839
- Biosynthetic (acetoacetyl-CoA-forming) activity: PMID:25478839
- Active-site residues: Cys92 (acyl-thioester intermediate nucleophile) and Cys382 (proton donor/acceptor); mutagenesis of C92 and C382 decreased acyl-CoA hydrolase activity (UniProt FT MUTAGEN).
- Kinetics (UniProt, from PMID:25478839): KM 9.2 uM acetoacetyl-CoA; KM 250 uM acetyl-CoA; KM 35 uM octanoyl-CoA; kcat 14.8 s^-1 for acetoacetyl-CoA degradation, 1.4 s^-1 for acetoacetyl-CoA synthesis, 0.02 s^-1 for octanoyl-CoA hydrolysis (hydrolysis is a very slow side reaction).
This is the EXP/IDA basis for the GOA molecular-function annotations:
GO:0003988 acetyl-CoA C-acyltransferase activity (EC 2.3.1.16, the core thiolase),
GO:0003985 acetyl-CoA C-acetyltransferase activity (EC 2.3.1.9, acetoacetyl-CoA reaction),
GO:0003986 acetyl-CoA hydrolase activity and GO:0047617 fatty acyl-CoA hydrolase activity
(the slow thioesterase side reaction).
ACAA2 was identified as a BNIP3-binding partner in a membrane yeast two-hybrid screen:
- PMID:18371312
- PMID:18371312
- PMID:18371312
- PMID:18371312
This is the basis for the BHF-UCL/UniProt process annotations: GO:0005515 protein binding
(IPI with BNIP3 = UniProtKB:Q12983), GO:0071456 cellular response to hypoxia,
GO:1902109 / GO:1901029 negative regulation of mitochondrial membrane permeability /
outer membrane permeabilization in apoptosis, and GO:0005739 mitochondrion (IDA).
Note: this is an overexpression/co-IP study; the apoptosis-modulation role is a
secondary/contextual function (single lab, overexpression-based), not the core
catalytic function. Should be KEPT but as non-core where applicable.
High-throughput mRNA-interactome capture in HeLa cells. ACAA2 was among proteins
crosslinked to poly(A) RNA:
- PMID:22658674
This is a proteome-wide HDA "moonlighting"-type detection, no gene-specific functional
follow-up for ACAA2. GO:0003723 RNA binding rests on an orthogonal high-throughput
screen; keep as non-core (not a characterized ACAA2 function).
Original cDNA cloning paper:
- PMID:8241273
The abstract describes only cloning/sequencing and Northern expression in liver,
fibroblasts, muscle. It does NOT mention cholesterol biosynthesis. The NAS
GO:0006695 cholesterol biosynthetic process annotation is not supported by this
abstract and is biologically implausible for a mitochondrial beta-oxidation thiolase
(cholesterol synthesis uses the cytosolic acetoacetyl-CoA thiolase ACAT2/cytosolic
HMG-CoA pathway, not mitochondrial ACAA2). Candidate REMOVE.
High-confidence human mitochondrial proteome study; HTP localization of ACAA2 to
mitochondrion. Corroborates mitochondrial localization. ACCEPT (localization).
CORE (ACCEPT, molecular function):
- GO:0003988 acetyl-CoA C-acyltransferase activity (EC 2.3.1.16) — IDA PMID:25478839 = THE core MF.
- GO:0006635 fatty acid beta-oxidation — core BP.
- mitochondrial matrix / mitochondrion — core localization.
ACCEPT non-core / supporting MF:
- GO:0003985 acetyl-CoA C-acetyltransferase activity (EC 2.3.1.9) — demonstrated biosynthetic activity, secondary.
- GO:0003986 acetyl-CoA hydrolase / GO:0047617 fatty acyl-CoA hydrolase — slow in-vitro side activity (kcat 0.02/s); KEEP_AS_NON_CORE.
IEA generic acyltransferase (GO:0016746, GO:0016747): less informative parents of the
specific thiolase MF; MARK_AS_OVER_ANNOTATED / generalize (MODIFY to GO:0003988) — keep as ACCEPT-but-general; use MARK_AS_OVER_ANNOTATED since covered by specific term.
RHEA hydrolase chain-length terms (GO:0052815 medium-chain, GO:0052816 long-chain,
GO:0141126 short-chain fatty acyl-CoA hydrolase): auto from Rhea mapping of the in-vitro
thioesterase side reactions. Physiologically minor (very low kcat). KEEP_AS_NON_CORE.
REMOVE candidates:
- GO:0006695 cholesterol biosynthetic process (NAS PMID:8241273) — unsupported by the
cited abstract, biologically wrong branch for mito beta-oxidation thiolase.
protein binding GO:0005515 (IPI BNIP3): do not endorse as core; KEEP_AS_NON_CORE
(documents real BNIP3 interaction but uninformative MF term).