Carnitine O-acetyltransferase

 

Carnitine acetyltransferase (CrAT) belongs to the family of choline/carnitine acyltransferases, which catalyse the exchange between acyl-CoA and acylcarnitine (likewise with choline). The enzymes of this family are classified based on their substrate selectivity. CrAT prefers short-chain fatty acids. CrAT maybe involved in the transport of acetyl-CoA across intercellular membranes, in maintaining the acetyl-CoA:CoA balance and in the excretion of excess or harmful acyl molecules as acylcarnitines. Inherited deficiency of CrAT activity can lead to serious neurological and heart problems, and patients suffering from Alzhiemer's disease also have reduced CrAT activity.

 

Reference Protein and Structure

Sequence
P47934 UniProt (2.3.1.7, 2.3.1.137) IPR000542 (Sequence Homologues) (PDB Homologues)
Biological species
Mus musculus (house mouse) Uniprot
PDB
1ndi - Carnitine Acetyltransferase in complex with CoA (2.3 Å) PDBe PDBsum 1ndi
Catalytic CATH Domains
3.30.559.40 CATHdb 3.30.559.70 CATHdb (see all for 1ndi)
Click To Show Structure

Enzyme Reaction (EC:2.3.1.7)

acetyl-CoA(4-)
CHEBI:57288ChEBI
+
(R)-carnitine
CHEBI:16347ChEBI
→
O-acetyl-L-carnitine
CHEBI:57589ChEBI
+
coenzyme A(4-)
CHEBI:57287ChEBI
Alternative enzyme names: CATC, Acetyl-CoA-carnitine O-acetyltransferase, Acetylcarnitine transferase, Carnitine acetyl coenzyme A transferase, Carnitine acetylase, Carnitine acetyltransferase, Carnitine-acetyl-CoA transferase,

Enzyme Mechanism

Introduction

His343 acts as a general base to deprotonate the 3-hydroxyl group of the carnitine or the thiol group of CoA, depending on what is being acetylated. The activated hydroxyl or thiol group acts as a nucleophile to attack the carbonyl carbon in acyl-CoA or acylcarnitine. The oxyanion intermediate (with a negatively charged oxygen atom) is stabilised by Ser554, apart of the STS motif, which forms the oxyanion hole. The reaction mechanism also has the carnitine trimethylammonium group lowering the pKa around the attacking hydroxyl and also stabilising the oxyanion intermediate, an example of substrate-assisted catalysis. The same mechanism occurs with Choline O-Acetyltransferase

Catalytic Residues Roles

UniProt PDB* (1ndi)
Tyr107, Pro120 Tyr107(78)A, Pro120(91)A By analogy with choline O-acetyltransferase Tyr107 and Pro120 steric pack with His 334, which forces the His 334 to adopt a strained conformation with an intraresidue hydrogen bond between N-delta and its carbonyl oxygen. This is proposed to position N-epsilon and stabilise its non-protonated, nucleophilic state. This promotes it role as a general base in the first step of the reaction. steric role
His343 His343(314)A It acts as a base to deprotonate the hydroxyl group of carnitine or the thiol group of CoA to promote their nucleophilic attack on the carbonyl carbon of CoA or carnitine. hydrogen bond acceptor, proton acceptor, proton donor
Ser554 Ser554(525)A It stabilises the negatively charged intermediate by forming an oxyanion hole. hydrogen bond donor, electrostatic stabiliser
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, overall reactant used, intermediate formation, bimolecular nucleophilic addition, unimolecular elimination by the conjugate base, intermediate terminated, overall product formed, native state of enzyme regenerated

References

  1. Jogl G et al. (2003), Cell, 112, 113-122. Crystal Structure of Carnitine Acetyltransferase and Implications for the Catalytic Mechanism and Fatty Acid Transport. DOI:10.1016/s0092-8674(02)01228-x. PMID:12526798.
  2. Hsiao YS et al. (2006), J Biol Chem, 281, 28480-28487. Crystal structures of murine carnitine acetyltransferase in ternary complexes with its substrates. DOI:10.1074/jbc.M602622200. PMID:16870616.
  3. Kim AR et al. (2006), Biochemistry, 45, 14621-14631. Substrate binding and catalytic mechanism of human choline acetyltransferase. DOI:10.1021/bi061536l. PMID:17144655.
  4. Jogl G et al. (2004), Ann N Y Acad Sci, 1033, 17-29. Structure and function of carnitine acyltransferases. DOI:10.1196/annals.1320.002. PMID:15591000.
  5. Cai Y et al. (2004), EMBO J, 23, 2047-2058. Choline acetyltransferase structure reveals distribution of mutations that cause motor disorders. DOI:10.1038/sj.emboj.7600221. PMID:15131697.

Step 1. His343 deprotonates the 3'OH on carnitine.

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Catalytic Residues Roles

Residue Roles
His343(314)A hydrogen bond acceptor
Tyr107(78)A steric role
Pro120(91)A steric role
His343(314)A proton acceptor

Chemical Components

proton transfer, overall reactant used

Step 2. Activated 3'OH then performs a nucleophilic attack on acetyl-CoA to form a tetrahedral intermediate.

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Catalytic Residues Roles

Residue Roles
Ser554(525)A electrostatic stabiliser
Ser554(525)A hydrogen bond donor

Chemical Components

intermediate formation, overall reactant used, ingold: bimolecular nucleophilic addition

Step 3. Tetrahedral intermediate collapses.

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Catalytic Residues Roles

Residue Roles
Ser554(525)A electrostatic stabiliser, hydrogen bond donor

Chemical Components

ingold: unimolecular elimination by the conjugate base, intermediate terminated, overall product formed

Step 4. Co-A is reprotonated by His343.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His343(314)A proton donor

Chemical Components

proton transfer, overall product formed, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. His343 deprotonates the 3'OH on carnitine.

Step 2. Activated 3'OH then performs a nucleophilic attack on acetyl-CoA to form a tetrahedral intermediate.

Step 3. Tetrahedral intermediate collapses.

Step 4. Co-A is reprotonated by His343.

Products.

Contributors

Mei Leung, Gemma L. Holliday, Morwenna Hall, Steven Smith