Haloalkane dehalogenase (subfamily 2)

 

The Haloalkane dehalogenase LinB, found in bacteria, catalyses the convertion of a broad range of natural and synthetic halogenated compounds (e.g., alkanes, alkenes, cycloalkanes, alcohols, epoxides, carboxylic acids, esters, ethers, amides, and nitriles) by replacing the halogen atom (chloro, bromo, or iodo) with a hydroxyl group. It is of special interest in its ability to degrade the toxic compound 1,2,3,4,5,6-hexachlorocyclohexane. Thus study of the enzyme is important in understanding how bacteria are able to degrade xenobiotics and how pollutants can be detoxified using microorganisms. The enzyme is part of the alpha-beta hydrolase family and shows sequence and structural homology with a wide variety of haloalkane dehalogenases with different specificities.

 

Reference Protein and Structure

Sequence
D4Z2G1 UniProt (3.8.1.5) IPR023594 (Sequence Homologues) (PDB Homologues)
Biological species
Sphingobium japonicum UT26S (Bacteria) Uniprot
PDB
1cv2 - Hydrolytic haloalkane dehalogenase linb from sphingomonas paucimobilis UT26 AT 1.6 A resolution (1.58 Å) PDBe PDBsum 1cv2
Catalytic CATH Domains
3.40.50.1820 CATHdb (see all for 1cv2)
Click To Show Structure

Enzyme Reaction (EC:3.8.1.5)

water
CHEBI:15377ChEBI
+
1-haloalkane
CHEBI:18060ChEBI
→
halide anion
CHEBI:16042ChEBI
+
hydron
CHEBI:15378ChEBI
+
primary alcohol
CHEBI:15734ChEBI
Alternative enzyme names: 1-chlorohexane halidohydrolase, 1-haloalkane dehalogenase,

Enzyme Mechanism

Introduction

The haloalkane dehalogenases catalyze a replacement reaction in which a primary or secondary halogen of a small molecule substrate is replaced with a hydroxyl group with the associated release of inorganic halide and a proton. The catalytic cycle of these enzymes consists of following steps:

  1. substrate binding into a relatively hydrophobic pocket,
  2. nucleophilic substitution reaction of an aspartic acid residue on the carbon atom to which the halogen is bound, leading to formation of a halide ion and an alkyl-enzyme intermediate,
  3. a second nucleophilic substitution reaction (hydrolysis, by a histidine activated water) leading to the formation of an alcohol and a proton, and
  4. products (an alcohol, a halide, and a proton) release.
The individual steps of the catalytic cycle are balanced and the rate-limiting step can be different for individual enzyme–substrate pairs, ranging from limitation at the nucleophilic substitution, through hydrolysis of the alkyl-enzyme intermediate, to an alcohol or a halide ion release.

Catalytic Residues Roles

UniProt PDB* (1cv2)
Glu132 Glu132A Acts to modify the pKa of His 272 so that it remains in the correct protonation state for its role in catalysis. activator, electrostatic stabiliser
His272 His272A Acts as general acid base to deprotonate water, thus activating water so its lone pair can attack the covalent enzyme intermediate. activator, proton shuttle (general acid/base), electrostatic stabiliser
Asp108 Asp108A Acts as nucleophile on the electrophilic carbon atom to form a covalent enzyme intermediate which is hydrolysed to give the product. covalent catalysis
Asn38, Trp109 Asn38A, Trp109A Involved in stabilisation of the halogen, transition-states and product. 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

References

  1. Streltsov VA et al. (2003), Biochemistry, 42, 10104-10112. Haloalkane Dehalogenase LinB fromSphingomonas paucimobilisUT26:  X-ray Crystallographic Studies of Dehalogenation of Brominated Substrates†,‡. DOI:10.1021/bi027280a. PMID:12939138.
  2. Gross J et al. (2016), Chembiochem, 17, 1437-1441. Regio- and Enantioselective Sequential Dehalogenation ofrac-1,3-Dibromobutane by Haloalkane Dehalogenase LinB. DOI:10.1002/cbic.201600227. PMID:27223496.
  3. Hladilkova J et al. (2013), J Phys Chem B, 117, 14329-14335. Release of Halide Ions from the Buried Active Site of the Haloalkane Dehalogenase LinB Revealed by Stopped-Flow Fluorescence Analysis and Free Energy Calculations. DOI:10.1021/jp409040u. PMID:24151979.
  4. Heeb NV et al. (2012), Environ Sci Technol, 46, 6566-6574. Biotransformation of Hexabromocyclododecanes (HBCDs) with LinB—An HCH-Converting Bacterial Enzyme. DOI:10.1021/es2046487. PMID:22578084.
  5. Otyepka M et al. (2008), Proteins, 70, 707-717. Second step of hydrolytic dehalogenation in haloalkane dehalogenase investigated by QM/MM methods. DOI:10.1002/prot.21523. PMID:17729274.
  6. Kmunícek J et al. (2005), Biochemistry, 44, 3390-3401. Quantitative Analysis of Substrate Specificity of Haloalkane Dehalogenase LinB fromSphingomonas paucimobilisUT26†. DOI:10.1021/bi047912o. PMID:15736949.
  7. Oakley AJ et al. (2004), Biochemistry, 43, 870-878. Crystal Structure of Haloalkane Dehalogenase LinB fromSphingomonas paucimobilisUT26 at 0.95 Å Resolution:  Dynamics of Catalytic Residues†,‡. DOI:10.1021/bi034748g. PMID:14744129.
  8. Marek J et al. (2000), Biochemistry, 39, 14082-14086. Crystal Structure of the Haloalkane Dehalogenase fromSphingomonas paucimobilisUT26†,‡. DOI:10.1021/bi001539c.
  9. Marek J et al. (2000), Biochemistry, 56, 14082-14086. Crystal structure of the haloalkane dehalogenase fromSphingomonas paucimobilisUT26. DOI:10.1107/s0108767300025332. PMID:11087355.

Step 1.

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Catalytic Residues Roles

Residue Roles
Trp109A electrostatic stabiliser
Asn38A electrostatic stabiliser
Glu132A electrostatic stabiliser, activator
His272A electrostatic stabiliser, proton shuttle (general acid/base), activator
Asp108A covalent catalysis

Chemical Components

Step 1.

Contributors

Gemma L. Holliday, Peter Sarkies