Thermodynamic analysis of halide binding to haloalkane dehalogenase suggests the occurrence of large conformational changes

Thermodynamic analysis of halide binding to haloalkane dehalogenase suggests the occurrence of large conformational changes.

Haloalkane dehalogenase (DhlA) hydrolyzes short-chain haloalkanes to produce the corresponding alcohols and halide ions. Release of the halide ion from the active-site cavity can proceed via a two-step and a three-step route, which both contain slow enzyme isomerization steps. Thermodynamic analysis of bromide binding and release showed that the slow unimolecular isomerization steps in the thre...

Kinetic analysis and X-ray structure of haloalkane dehalogenase with a modified halide-binding site.

Haloalkane dehalogenase (DhlA) catalyzes the hydrolysis of haloalkanes via an alkyl-enzyme intermediate. Trp175 forms a halogen/halide-binding site in the active-site cavity together with Trp125. To get more insight in the role of Trp175 in DhlA, we mutated residue 175 and explored the kinetics and X-ray structure of the Trp175Tyr enzyme. The mutagenesis study indicated that an aromatic residue...

Kinetics of halide release of haloalkane dehalogenase: evidence for a slow conformational change.

Haloalkane dehalogenase converts haloalkanes to their corresponding alcohols and halides. The reaction mechanism involves the formation of a covalent alkyl-enzyme complex which is hydrolyzed by water. The active site is a hydrophobic cavity buried between the main domain and the cap domain of the enzyme. The enzyme has a broad substrate specificity, but the kcat values of the enzyme for the bes...

University of Groningen REPLACEMENT OF TRYPTOPHAN RESIDUES IN HALOALKANE DEHALOGENASE REDUCES HALIDE BINDING AND CATALYTIC ACTIVITY

Replacement of tryptophan residues in haloalkane dehalogenase reduces halide binding and catalytic activity.

Haloalkane dehalogenase catalyzes the hydrolytic cleavage of carbon-halogen bonds in short-chain haloalkanes. Two tryptophan residues of the enzyme (Trp125 and Trp175) form a halide-binding site in the active-site cavity, and were proposed to play a role in catalysis. The function of these residues was studied by replacing Trp125 with phenylalanine, glutamine or arginine and Trp175 by glutamine...