M-CSA Mechanism and Catalytic Site Atlas

Vanadium-dependent bromoperoxidase (HOBr Formation)

V-BPO, isolated from Ascophyllum nodosum, is a vanadium-dependent haloperoxidase. It catalyses the oxidation of halides by hydrogen peroxide to form hypohalides, which can go on to react with organic substrates in halogenation reactions. In this entry, we show the primary biological reaction of V-BPO: the formation of HOBr. V-BPO has the ability to oxidise bromide and, to a lesser extent, chloride and iodide as well as chlorinate organic hydrocarbons and oxidise organic sulfides. V-BPO contains a vanadium(V) ion with a trigonal bipyramidal coordination sphere. It is coordinated to His486 and a hydroxide in the axial positions, and two oxygen atoms and a hydroxide in the equatorial positions.

Reference Protein and Structure

Sequence: P81701 (1.11.1.18) (Sequence Homologues) (PDB Homologues)
Biological species: Ascophyllum nodosum (Knotted wrack)
PDB: 1qi9 - X-RAY SIRAS STRUCTURE DETERMINATION OF A VANADIUM-DEPENDENT HALOPEROXIDASE FROM ASCOPHYLLUM NODOSUM AT 2.0 A RESOLUTION (2.05 Å)
Catalytic CATH Domains: 1.10.606.10 (see all for 1qi9)
Cofactors: Vanadate(3-) (1)

Click To Show Structure

Enzyme Reaction (EC:1.11.1.18)

hydron

CHEBI:15378

bromide

CHEBI:15858

hydrogen peroxide

CHEBI:16240

→

water

CHEBI:15377

hypobromous acid

CHEBI:29249

Enzyme reaction links: IntEnz ENZYME ExplorEnz

Alternative enzyme names: Bromoperoxidase, Haloperoxidase, Eosinophil peroxidase,

Enzyme Mechanism

Mechanism Proposal 1 ☆☆☆

Summary
Step 1
Step 2
Step 3
Step 4
Step 5
Step 6
Products
All Steps

Introduction

An oxygen of hydrogen peroxide attacks the vanadium centre and causes the elimination of the axial hydroxide, which then deprotonates the bound peroxide to form water. The second oxygen of the peroxide ligand then attacks the vanadium centre and causes the elimination of an equatorial oxygen, which then abstracts the second peroxide proton. This forms a side-on vanadium-peroxo species. Bromide attacks one of the oxygens of the peroxo group, causing the O-O bond to break and leading to the formation of an axial vanadium-OBr complex. In the final step, the hypobromite ligand is protonated by water and is displaced by the resulting hydroxide ligand.

Catalytic Residues Roles

UniProt

PDB* (1qi9)

*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, bimolecular nucleophilic substitution, dehydration, intramolecular nucleophilic substitution, bimolecular nucleophilic addition

References

Weyand M et al. (1999), J Mol Biol, 293, 595-611. X-ray structure determination of a vanadium-dependent haloperoxidase from Ascophyllum nodosum at 2.0 Å resolution. DOI:10.1006/jmbi.1999.3179. PMID:10543953.
Waller MP et al. (2008), J Phys Chem B, 112, 5813-5823. 51V NMR Chemical Shifts from Quantum-Mechanical/Molecular-Mechanical Models of Vanadium Bromoperoxidase. DOI:10.1021/jp800580n. PMID:18412416.
Raugei S et al. (2006), J Phys Chem B, 110, 3747-3758. Structure and Function of Vanadium Haloperoxidases†. DOI:10.1021/jp054901b. PMID:16494433.
Zampella G et al. (2005), J Am Chem Soc, 127, 953-960. Reactivity of Peroxo Forms of the Vanadium Haloperoxidase Cofactor. A DFT Investigation. DOI:10.1021/ja046016x. PMID:15656634.
Rehder D et al. (2000), J Inorg Biochem, 80, 115-121. Water and bromide in the active center of vanadate-dependent haloperoxidases. DOI:10.1016/s0162-0134(00)00047-7. PMID:10885471.
Macedo-Ribeiro S et al. (1999), J Biol Inorg Chem, 4, 209-219. X-ray crystal structures of active site mutants of the vanadium-containing chloroperoxidase from the fungus Curvularia inaequalis. DOI:10.1007/s007750050306. PMID:10499093.

Step 1. The axial hydroxide of the vanadate cofactor deprotonates hydrogen peroxide.

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Catalytic Residues Roles

Residue	Roles
His486A	metal ligand, electrostatic stabiliser
Arg349A	electrostatic stabiliser
His411A	increase basicity
His418A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator
Lys341A	electrostatic stabiliser

Chemical Components

proton transfer

Step 2. The activated hydrogen peroxide initiates a nucleophilic attack on the vanadate in a substitution reaction, eliminating water.

Download: Image, Marvin File

Catalytic Residues Roles

Residue	Roles
His486A	metal ligand, activator
Arg349A	electrostatic stabiliser
Lys341A	electrostatic stabiliser
His411A	electrostatic stabiliser
His418A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator

Chemical Components

ingold: bimolecular nucleophilic substitution, dehydration

Step 3. One of the equatorial oxo groups deprotonates the attached hydrogen peroxide.

Download: Image, Marvin File

Catalytic Residues Roles

Residue	Roles
His486A	metal ligand, activator
Arg349A	electrostatic stabiliser
Lys341A	electrostatic stabiliser
His418A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator

Chemical Components

proton transfer

Step 4. The peroxide initiates a nucleophilic attack on the vanadate in a substitution reaction, eliminating hydroxide and forming a three membered ring.

Download: Image, Marvin File

Catalytic Residues Roles

Residue	Roles
His486A	metal ligand, activator
Lys341A	electrostatic stabiliser
His418A	electrostatic stabiliser
Arg349A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator

Chemical Components

ingold: intramolecular nucleophilic substitution

Step 5. The halide adds to one of the peroxide oxygen atoms.

Download: Image, Marvin File

Catalytic Residues Roles

Residue	Roles
His418A	electrostatic stabiliser
His486A	metal ligand, activator
Lys341A	electrostatic stabiliser
Arg349A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator

Chemical Components

ingold: bimolecular nucleophilic addition

Step 6. Hypobromate is eliminated with concomitant deprotonation of water, which initiates a nucleophilic attack on the vanadate in a coordination reaction to regenerate the cofactor.

Download: Image, Marvin File

Catalytic Residues Roles

Residue	Roles
His418A	electrostatic stabiliser
His486A	metal ligand, activator
Lys341A	electrostatic stabiliser
His411A	increase acidity
Arg349A	electrostatic stabiliser
Ser416A	electrostatic stabiliser
Gly417A (main-N)	electrostatic stabiliser
Arg480A	electrostatic stabiliser
Asp490A	activator

Chemical Components

proton transfer, ingold: bimolecular nucleophilic substitution

Download: Image, Marvin File

Step 1. The axial hydroxide of the vanadate cofactor deprotonates hydrogen peroxide.

Step 2. The activated hydrogen peroxide initiates a nucleophilic attack on the vanadate in a substitution reaction, eliminating water.

Step 3. One of the equatorial oxo groups deprotonates the attached hydrogen peroxide.

Step 4. The peroxide initiates a nucleophilic attack on the vanadate in a substitution reaction, eliminating hydroxide and forming a three membered ring.

Step 5. The halide adds to one of the peroxide oxygen atoms.

Step 6. Hypobromate is eliminated with concomitant deprotonation of water, which initiates a nucleophilic attack on the vanadate in a coordination reaction to regenerate the cofactor.

Products.

Contributors

Gemma L. Holliday