Malonyl-CoA-acyl carrier protein transacylase

 

In Escherichia coli Fatty acid synthesis, FAS II includes a specific malonyl-CoA:ACP transacylase (MCAT), which catalyses specifically the elongation step. The initiation of each elongation step in the fatty acid synthesis cycle requires the transfer of a malonyl moiety from the respective CoA thioester to the -SH group of the phosphopantetheine arm of the acyl carrier protein (ACP), the central component of any FAS. MCAT, along with Acyl-carrier-protein S-acetyltransferase (EC 2.3.1.38), is essential for the initiation of fatty-acid biosynthesis in bacteria. This enzyme also provides the malonyl groups for polyketide biosynthesis.

 

Reference Protein and Structure

Sequence
P0AAI9 UniProt (2.3.1.39) IPR004410 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1mla - THE ESCHERICHIA COLI MALONYL-COA:ACYL CARRIER PROTEIN TRANSACYLASE AT 1.5-ANGSTROMS RESOLUTION. CRYSTAL STRUCTURE OF A FATTY ACID SYNTHASE COMPONENT (1.5 Å) PDBe PDBsum 1mla
Catalytic CATH Domains
3.40.366.10 CATHdb (see all for 1mla)
Click To Show Structure

Enzyme Reaction (EC:2.3.1.39)

O-(pantetheine-4'-phosphoryl)serine(1-) residue
CHEBI:64479ChEBI
+
malonyl-CoA(5-)
CHEBI:57384ChEBI
→
coenzyme A(4-)
CHEBI:57287ChEBI
+
O-(S-malonylpantetheine-4'-phosphoryl)serine(2-) residue
CHEBI:78449ChEBI
Alternative enzyme names: [Acyl carrier protein]malonyltransferase, Malonyl coenzyme A-acyl carrier protein transacylase, Malonyl transacylase, Malonyl transferase, Malonyl-CoA-acyl carrier protein transacylase, MAT, FabD, Malonyl-CoA:acyl carrier protein transacylase, Malonyl-CoA:ACP transacylase, MCAT, Malonyl-CoA:AcpM transacylase, Acyl carrier protein malonyltransferase, Malonyl-CoA:[acyl-carrier-protein] S-malonyltransferase, Malonyl-CoA:ACP-SH transacylase, Malonyl-CoA:acyl-carrier-protein transacylase, Malonyl-CoA/dephospho-CoA acyltransferase, MdcH,

Enzyme Mechanism

Introduction

The catalytic residues consist of a conventional Ser-His dyad, which is unusually hydrogen to the main chain carbonyl of a glutamine residue. The reactions consists of an acylation step and the subsequent transfer of the acyl moiety proceed via tetrahedral intermediates, analogously to serine proteases. His201 deprotonates Ser92 to form a nucleophile. This subsequently attacks the carbonyl group-subsequent break down of this tetrahedral intermediate eliminates CoA forming a thioester with the enzyme. His201 then deprotonates a water molecule, which nucleophilically attacks the carbonyl group a second time break down results in elimination of the enzyme.

Catalytic Residues Roles

UniProt PDB* (1mla)
Ser92 Ser92A Deprotonated by His201, performs a nucleophilic attack on the carbonyl group of the substrate to form a tetrahedral intermediate. covalently attached, hydrogen bond acceptor, hydrogen bond donor, nucleophile, proton acceptor, proton donor, nucleofuge, activator
His201 His201A Deprotonates Ser92 to activate it for nucleophilic attack. Similarly deprotonates a water molecule. Acts as a general acid to leaving groups of both tetrahedral intermediates. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor, increase nucleophilicity
Leu93 (main-N), Arg117, Gln11 Leu93A (main-N), Arg117A, Gln11A Act to stabilise the reactive intermediates and transition states formed during the course of the reaction. hydrogen bond donor, electrostatic stabiliser
Gln250 (main-C) Gln250A (main-C) Forms a hydrogen bond to His201 to aid it in its role as general acid/base. increase basicity, hydrogen bond acceptor, 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

bimolecular nucleophilic substitution, proton transfer, enzyme-substrate complex formation, intermediate formation, overall reactant used, unimolecular elimination by the conjugate base, intermediate collapse, enzyme-substrate complex cleavage, overall product formed, bimolecular nucleophilic addition, native state of enzyme regenerated

References

  1. Serre L et al. (1995), J Biol Chem, 270, 12961-12964. The Escherichia coli malonyl-CoA:acyl carrier protein transacylase at 1.5-A resolution. Crystal structure of a fatty acid synthase component. DOI:10.2210/pdb1mla/pdb. PMID:7768883.
  2. Paiva P et al. (2018), ACS Catal, 8, 4860-4872. Understanding the Catalytic Machinery and the Reaction Pathway of the Malonyl-Acetyl Transferase Domain of Human Fatty Acid Synthase. DOI:10.1021/acscatal.8b00577.
  3. Serre L et al. (1994), J Mol Biol, 242, 99-102. Crystallization of the Malonyl Coenzyme A-Acyl Carrier Protein Transacylase from Escherichia coli. DOI:10.1006/jmbi.1994.1559. PMID:8078074.
  4. Joshi VC (1972), Biochem J, 128, 43P.2-44P. Mechanism of malonyl-coenzyme A-acyl-carrier protein transacylase. DOI:10.1042/bj1280043pb. PMID:4563767.

Step 1. Ser92 is activated towards nucleophilic attack at the thio-carbonyl of malonyl-CoA by the general base His201. A catalytic dyad is present between Ser92A and His201A, with the protonated form of His201A being stabilised through a hydrogen bond with the main chain carbonyl of Gln250A [PMID:7768883].

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Catalytic Residues Roles

Residue Roles
Ser92A hydrogen bond donor
His201A hydrogen bond acceptor, increase nucleophilicity
Gln250A (main-C) hydrogen bond acceptor, increase basicity
Gln11A hydrogen bond donor, electrostatic stabiliser
Arg117A hydrogen bond donor, electrostatic stabiliser, attractive charge-charge interaction
Ser92A proton donor
His201A proton acceptor
Ser92A nucleophile

Chemical Components

ingold: bimolecular nucleophilic substitution, proton transfer, enzyme-substrate complex formation, intermediate formation, overall reactant used

Step 2. The tetrahedral enzyme-substrate intermediate collapses, eliminating the coenzyme A component.

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Catalytic Residues Roles

Residue Roles
Ser92A covalently attached, activator
His201A hydrogen bond donor
Gln250A (main-C) hydrogen bond acceptor, electrostatic stabiliser
Gln11A hydrogen bond donor, electrostatic stabiliser
Arg117A hydrogen bond donor, attractive charge-charge interaction, electrostatic stabiliser
Leu93A (main-N) hydrogen bond donor, electrostatic stabiliser

Chemical Components

ingold: unimolecular elimination by the conjugate base, intermediate collapse, intermediate formation, enzyme-substrate complex cleavage

Step 3. The terminal thiolate of the previously eliminated CoA acts as a general base towards His201.

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Catalytic Residues Roles

Residue Roles
Ser92A covalently attached
His201A hydrogen bond donor
Gln250A (main-C) hydrogen bond acceptor, electrostatic stabiliser
Gln11A hydrogen bond donor
Arg117A hydrogen bond donor
Leu93A (main-N) hydrogen bond donor
His201A proton donor

Chemical Components

proton transfer, overall product formed

Step 4. The enzyme-malonyl carbonyl linkage acts as an electrophile towards the terminal thiol of the acyl carrier protein which is deprotonated by the general acid His201.

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Catalytic Residues Roles

Residue Roles
His201A proton acceptor

Chemical Components

ingold: bimolecular nucleophilic addition, proton transfer, intermediate formation, overall reactant used, enzyme-substrate complex formation

Step 5. The tetrahedral intermediate collapses eliminating the malonyl acyl-carrier protein adduct and free serine side chain, which is reprotonated by the general acid His201A.

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Catalytic Residues Roles

Residue Roles
Ser92A covalently attached, hydrogen bond acceptor
His201A hydrogen bond donor
Gln11A hydrogen bond donor
Arg117A hydrogen bond donor
His201A proton donor
Ser92A proton acceptor, nucleofuge

Chemical Components

proton transfer, ingold: unimolecular elimination by the conjugate base, intermediate collapse, enzyme-substrate complex cleavage, overall product formed, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. Ser92 is activated towards nucleophilic attack at the thio-carbonyl of malonyl-CoA by the general base His201. A catalytic dyad is present between Ser92A and His201A, with the protonated form of His201A being stabilised through a hydrogen bond with the main chain carbonyl of Gln250A [PMID:7768883].

Step 2. The tetrahedral enzyme-substrate intermediate collapses, eliminating the coenzyme A component.

Step 3. The terminal thiolate of the previously eliminated CoA acts as a general base towards His201.

Step 4. The enzyme-malonyl carbonyl linkage acts as an electrophile towards the terminal thiol of the acyl carrier protein which is deprotonated by the general acid His201.

Step 5. The tetrahedral intermediate collapses eliminating the malonyl acyl-carrier protein adduct and free serine side chain, which is reprotonated by the general acid His201A.

Products.

Contributors

Sophie T. Williams, Gemma L. Holliday, Alex Gutteridge, Craig Porter, Charity Hornby