7-hydroxymethyl chlorophyll A reductase (HCAR)

 

7-Hydroxymethyl chlorophyll (HCAR) is a member of the family of chlorophyll catabolic enzymes (CCEs). Its a reductase that catalyses one of the reactions in chlorophyll conversion. HCAR is required for the degradation of light-harvesting complexes and is necessary for efficient photosynthesis by balancing the chlorophyll A/B ratio. Specifically, its a protein containing both 4Fe-4S complexes and FAD cofactor that converts 7-hydroxymethyl chlorophyll (HMChl) a to chlorophyll A using ferredoxin for reduction of the substrate. HCAR catalyses the reduction of a hydroxymethyl group to a methyl group.

 

Reference Protein and Structure

Sequence
Q8GS60 UniProt (1.17.7.2) IPR007525 (Sequence Homologues) (PDB Homologues)
Biological species
Arabidopsis thaliana (Thale cress) Uniprot
PDB
5dqr - The crystal structure of Arabidopsis 7-hydroxymethyl chlorophyll a reductase (HCAR) (2.7 Å) PDBe PDBsum 5dqr
Catalytic CATH Domains
(see all for 5dqr)
Click To Show Structure

Enzyme Reaction (EC:1.17.7.2)

7(1)-hydroxychlorophyll a(1-)
CHEBI:83377ChEBI
+
hydron
CHEBI:15378ChEBI
+
di-mu-sulfido-diiron(1+)
CHEBI:33738ChEBI
→
chlorophyll a(1-)
CHEBI:58416ChEBI
+
water
CHEBI:15377ChEBI
+
di-mu-sulfido-diiron(2+)
CHEBI:33737ChEBI
Alternative enzyme names: HCAR, 7(1)-hydroxychlorophyll a:ferredoxin oxidoreductase,

Enzyme Mechanism

Introduction

The 7-hydroxychlorophyll (HMChl) to chlorophyll A (ChlA) reaction is done by a mechanism in which protons coming from Asp237 and solution and electrons coming from ferrodoxin, through 2 4Fe-4S molecules and to an FAD molecule, are able to dehydrate and reduce the HMChl. Asp237 donates a proton to dehydrate the HMChl, this causes bond rearrangement in the chlorophyll structure and the formation of a double bond on the C atom that lost its hydroxyl group. The electron transfer then begins with an external ferrodoxin protein donating 2 electrons. These electrons pass over two 4Fe-4S molecules onto an FAD cofactor. FAD then donates these electrons to the methenyl group on the substrate, resulting in a methyl group.

Catalytic Residues Roles

UniProt PDB* (5dqr)
Asp237 Asp237(212)D Proton donor crucial for dehydrating the HMChl. proton acceptor, proton donor
His417 His417(392)D Stabilises and binds the HMChl molecule. metal ligand, electrostatic stabiliser
Glu262, Gln332 Glu262(237)D, Gln332(307)D Glu262 and Gln332 stabilise and anchor FAD. Mutations on these residues lead to complete loss of function. 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

dehydration, intramolecular rearrangement, unimolecular elimination by the conjugate base, radical formation, electron transfer, electron relay, proton transfer, inferred reaction step, radical termination, hydride transfer, redox reaction, overall product formed, overall reactant used

References

  1. Wang X et al. (2016), J Biol Chem, 291, 13349-13359. Crystal Structure and Catalytic Mechanism of 7-Hydroxymethyl Chlorophyll a Reductase. DOI:10.1074/jbc.M116.720342. PMID:27072131.
  2. Meguro M et al. (2011), Plant Cell, 23, 3442-3453. Identification of the 7-Hydroxymethyl Chlorophyll a Reductase of the Chlorophyll Cycle in Arabidopsis  . DOI:https://doi.org/10.1105/tpc.111.089714.

Step 1. Asp237 protonates the OH group on the 7-hydroxychlorophyll a (HMChl). This causes a water molecule to leave HMChl. The substrate then goes through bond rearrangement and creates a double bond towards the carbon atom that lost its hydroxyl group to stabilise the molecule.

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Catalytic Residues Roles

Residue Roles
His417(392)D electrostatic stabiliser, metal ligand
Asp237(212)D proton donor

Chemical Components

dehydration, intramolecular rearrangement, ingold: unimolecular elimination by the conjugate base

Step 2. Ferrodoxin donates a electron to the 4Fe-4S molecules which relay them to FAD. This process is stabilised by Gln332 and Glu262. The protonation from the bulk solvent is inferred.

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Catalytic Residues Roles

Residue Roles
Glu262(237)D electrostatic stabiliser
Gln332(307)D electrostatic stabiliser
His417(392)D electrostatic stabiliser

Chemical Components

radical formation, intramolecular rearrangement, electron transfer, electron relay, proton transfer, inferred reaction step

Step 3. Ferrodoxin donates a second electron which is used to fully reduce FAD. Another proton from solution is used to stabilise the negative charge.

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Catalytic Residues Roles

Residue Roles
Glu262(237)D electrostatic stabiliser
Gln332(307)D electrostatic stabiliser
His417(392)D electrostatic stabiliser, metal ligand

Chemical Components

radical termination, electron relay, electron transfer, proton transfer, inferred reaction step

Step 4. The electrons gained by FAD are given to the chlorophyll substrate in the form of a hydride transfer. Asp237 is regenerated to its protonated state. The chlorophyll substrate undergoes some bond rearrangements to stabilise the added electrons.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp237(212)D proton acceptor
His417(392)D electrostatic stabiliser
Glu262(237)D electrostatic stabiliser
Gln332(307)D electrostatic stabiliser
His417(392)D metal ligand

Chemical Components

hydride transfer, intramolecular rearrangement, proton transfer, redox reaction, overall product formed, overall reactant used
Download: Image, Marvin File

Step 1. Asp237 protonates the OH group on the 7-hydroxychlorophyll a (HMChl). This causes a water molecule to leave HMChl. The substrate then goes through bond rearrangement and creates a double bond towards the carbon atom that lost its hydroxyl group to stabilise the molecule.

Step 2. Ferrodoxin donates a electron to the 4Fe-4S molecules which relay them to FAD. This process is stabilised by Gln332 and Glu262. The protonation from the bulk solvent is inferred.

Step 3. Ferrodoxin donates a second electron which is used to fully reduce FAD. Another proton from solution is used to stabilise the negative charge.

Step 4. The electrons gained by FAD are given to the chlorophyll substrate in the form of a hydride transfer. Asp237 is regenerated to its protonated state. The chlorophyll substrate undergoes some bond rearrangements to stabilise the added electrons.

Products.

Contributors

Marko Babić, Antonio Ribeiro