Cutinase

 

Cutinase is a serine esterase. It hydrolyses cutin, an insoluble polyester which covers the surface of plants. It is also able to hydrolyse fatty acids esters and emulsified triacylglycerol as efficiently as lipases.

 

Reference Protein and Structure

Sequence
P00590 UniProt (3.1.1.74) IPR011150 (Sequence Homologues) (PDB Homologues)
Biological species
Fusarium vanettenii Uniprot
PDB
1agy - The 1.15 angstrom refined structure of fusarium solani pisi cutinase (1.15 Å) PDBe PDBsum 1agy
Catalytic CATH Domains
3.40.50.1820 CATHdb (see all for 1agy)
Click To Show Structure

Enzyme Reaction (EC:3.1.1.74)

water
CHEBI:15377ChEBI
+
carboxylic ester
CHEBI:33308ChEBI
→
alcohol
CHEBI:30879ChEBI
+
carboxylic acid anion
CHEBI:29067ChEBI
+
hydron
CHEBI:15378ChEBI

Enzyme Mechanism

Introduction

His204 acts as a base to deprotonate Ser136 to allow its nucleophilic attack on the ester bond. His204 donates a proton to the leaving group and then activates a water molecule to allow the hydrolysis of the acylenzyme. Asp191 alters the pKa of the His204 to allow to act as an effective base in the reaction. Gln137 and Ser58 forms the oxyanion hole to stabilise the transition state.

Catalytic Residues Roles

UniProt PDB* (1agy)
His204 His188(174)A It deprotonates Ser 136 to allow its nucleophilic attack on the ester bond. It donates a proton to the leaving group. It activates a water molecule to promote the hydrolysis of the acylenzyme intermediate. proton acceptor, proton donor
Asp191 Asp175(161)A It alters the pKa of His 204 to allow it to act as an effective acid/base in the reaction. increase basicity, electrostatic stabiliser
Ser136 Ser120(106)A It acts as a nucleophile to attack the ester bond. covalently attached, nucleofuge, nucleophile, proton acceptor, proton donor
Ser58 (main-N), Gln137 (main-N) Ser42(28)A (main-N), Gln121(107)A (main-N) It forms the oxyanion hole, stabilising the tetrahedral transition state. 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 addition, proton transfer, intermediate formation, overall reactant used, unimolecular elimination by the conjugate base, overall product formed, intermediate terminated, native state of enzyme regenerated

References

  1. Martinez C et al. (1994), Biochemistry, 33, 83-89. Cutinase, a lipolytic enzyme with a preformed oxyanion hole. DOI:10.1021/bi00167a011. PMID:8286366.
  2. Nyon MP et al. (2009), J Mol Biol, 385, 226-235. Catalysis by Glomerella cingulata cutinase requires conformational cycling between the active and inactive states of its catalytic triad. DOI:10.1016/j.jmb.2008.10.050. PMID:18983850.
  3. Martinez C et al. (1992), Nature, 356, 615-618. Fusarium solani cutinase is a lipolytic enzyme with a catalytic serine accessible to solvent. DOI:10.1038/356615a0. PMID:1560844.

Step 1. His204 acts as a general base activating the Ser136 hydroxyl group for nucleophilic attack on the carbonyl carbon. The third component of this catalytic triad- Asp191 acts to increase the basicity of the histidine. The oxyanion intermediate formed is stabilized by the amide groups of Gln137 and Ser58.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Ser42(28)A (main-N) electrostatic stabiliser
Gln121(107)A (main-N) electrostatic stabiliser
Asp175(161)A electrostatic stabiliser
Ser120(106)A covalently attached
Asp175(161)A increase basicity
Ser120(106)A nucleophile
His188(174)A proton acceptor
Ser120(106)A proton donor

Chemical Components

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

Step 2. The tetrahedral intermediate collapses and an alcohol is eliminated.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Ser42(28)A (main-N) electrostatic stabiliser
Gln121(107)A (main-N) electrostatic stabiliser
Asp175(161)A electrostatic stabiliser
Ser120(106)A covalently attached
His188(174)A proton donor

Chemical Components

ingold: unimolecular elimination by the conjugate base, proton transfer, overall product formed

Step 3. His204 activates water for nucleophilic attack and another oxyanion intermediate is formed.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Ser42(28)A (main-N) electrostatic stabiliser
Gln121(107)A (main-N) electrostatic stabiliser
Asp175(161)A electrostatic stabiliser, increase basicity
Ser120(106)A covalently attached
His188(174)A proton acceptor

Chemical Components

proton transfer, ingold: bimolecular nucleophilic addition

Step 4. The tetrahedral intermediate collapses and Ser136 is eliminated.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Ser42(28)A (main-N) electrostatic stabiliser
Gln121(107)A (main-N) electrostatic stabiliser
Asp175(161)A electrostatic stabiliser
Ser120(106)A proton acceptor, nucleofuge
His188(174)A proton donor

Chemical Components

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

Step 1. His204 acts as a general base activating the Ser136 hydroxyl group for nucleophilic attack on the carbonyl carbon. The third component of this catalytic triad- Asp191 acts to increase the basicity of the histidine. The oxyanion intermediate formed is stabilized by the amide groups of Gln137 and Ser58.

Step 2. The tetrahedral intermediate collapses and an alcohol is eliminated.

Step 3. His204 activates water for nucleophilic attack and another oxyanion intermediate is formed.

Step 4. The tetrahedral intermediate collapses and Ser136 is eliminated.

Products.

Contributors

Mei Leung, Gemma L. Holliday, James Willey