Carboxy-cis,cis-muconate cyclase

 

Eukaryote 3-carboxy-cis,cis-muconate lactonizing enzyme is an eukaryote muconate lactonizing enzyme(MLE), converting cis,cis-muconates to muconlactones. It is in the class of CyclicMLE among the three classes of muconate lactonizing enzymes. The other 2 classes are bacterial CyclicMLE and bacterial MLE. The three classes of MLE are structurally very different but catalyse an analogous reaction, i.e. convergent functional evolution occurred.

 

Reference Protein and Structure

Sequence
P38677 UniProt (5.5.1.5) IPR019405 (Sequence Homologues) (PDB Homologues)
Biological species
Neurospora crassa OR74A (Fungus) Uniprot
PDB
1jof - Neurospora crassa 3-carboxy-cis,cis-mucoante lactonizing enzyme (2.5 Å) PDBe PDBsum 1jof
Catalytic CATH Domains
2.130.10.10 CATHdb (see all for 1jof)
Click To Show Structure

Enzyme Reaction (EC:5.5.1.5)

2-(carboxylatomethyl)-5-oxo-2,5-dihydrofuran-2-ide-3-carboxylate
CHEBI:57976ChEBI
→
3-carboxy-cis,cis-muconate(3-)
CHEBI:57496ChEBI
Alternative enzyme names: 3-carboxymuconate cyclase, 3-carboxy-cis,cis-muconate lactonizing enzyme, 3-carboxy-2,5-dihydro-5-oxofuran-2-acetate lyase (decyclizing),

Enzyme Mechanism

Introduction

The mechanism is a general acid/base catalysis, where His 148 deprotonates C5 and Glu 212 donates a proton to the C6 carboxylate. This causes a tautomerization to take place forming an eneolate intermediate, stabilized by Arg 274 and 196 via hydrogen bonding. Intramolecular nucleophilic attack from the C6 enolate to C4 via the C=C bond causes the enolate to tautomerize back into a carboxylate and for the 5 member ring to be opened, forming another carboxylate group on C1. The Arg residues act to increase nucleophilicity of the enolate via hydrogen bonding, allowing the product to be formed in this step.

Catalytic Residues Roles

UniProt PDB* (1jof)
Arg197 Arg196A Hydrogen bonded to the substrate C6 carboxylate group oxygen to increase the nucleophilicity of the group increase nucleophilicity, electrostatic stabiliser
Glu213 Glu212A Donates a proton to the C6 carboxylate in the initial tautomerization step. proton donor
Arg275 Arg274A Hydrogen bonded to the substrate C6 carboxylate group oxygen to increase its nucleophilicity. increase nucleophilicity, electrostatic stabiliser
His149 His148A Acts as a base to deprotonate C5 in the initial tautomerization. proton acceptor
*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, assisted keto-enol tautomerisation, overall reactant used, rate-determining step, keto-enol tautomerisation, intramolecular nucleophilic substitution, overall product formed

References

  1. Kajander T et al. (2002), Structure, 10, 483-492. The Structure of Neurospora crassa 3-Carboxy-cis,cis-Muconate Lactonizing Enzyme, a β Propeller Cycloisomerase. DOI:10.1016/s0969-2126(02)00744-x. PMID:11937053.
  2. Yang J et al. (2004), Biochemistry, 43, 10424-10434. Crystal structure of 3-carboxy-cis,cis-muconate lactonizing enzyme from Pseudomonas putida, a fumarase class II type cycloisomerase: enzyme evolution in parallel pathways. DOI:10.1021/bi036205c. PMID:15301541.

Step 1. His 148 deprotonates C5, simultaneously Glu 212 donates a proton to the C6 carboxylate. This causes a tautomerization to take place forming an eneolate which is stabilized by the Arg residues via hydrogen bonding.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Arg274A electrostatic stabiliser
Arg196A electrostatic stabiliser
His148A proton acceptor
Glu212A proton donor

Chemical Components

proton transfer, assisted keto-enol tautomerisation, overall reactant used, rate-determining step

Step 2. Intramolecular nucleophilic attack from the C6 enolate to C4 via the C=C bond causes the enolate to tautomerize back into a carboxylate and for the 5 member ring to be opened, forming another carboxylate group on C1. The Arg residues act to increase nucleophilicity of the enolate via hydrogen bonding.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Arg196A increase nucleophilicity
Arg274A increase nucleophilicity
Arg196A electrostatic stabiliser
Arg274A electrostatic stabiliser

Chemical Components

keto-enol tautomerisation, ingold: intramolecular nucleophilic substitution, overall product formed
Download: Image, Marvin File

Step 1. His 148 deprotonates C5, simultaneously Glu 212 donates a proton to the C6 carboxylate. This causes a tautomerization to take place forming an eneolate which is stabilized by the Arg residues via hydrogen bonding.

Step 2. Intramolecular nucleophilic attack from the C6 enolate to C4 via the C=C bond causes the enolate to tautomerize back into a carboxylate and for the 5 member ring to be opened, forming another carboxylate group on C1. The Arg residues act to increase nucleophilicity of the enolate via hydrogen bonding.

Products.

Contributors

Mei Leung, Gemma L. Holliday, James Willey