Protein engineering of the 2-haloacid halidohydrolase iva from pseudomonas-cepacia mab4

Asmara, W. and Murdiyatmo, Untung and Baines, Anthony J. and Bull, Alan T. and Hardman, David J. (1993) Protein engineering of the 2-haloacid halidohydrolase iva from pseudomonas-cepacia mab4. Biochemical Journal, 292 . pp. 69-74. ISSN 0264-6021. (The full text of this publication is not currently available from this repository. You may be able to access a copy if URLs are provided)

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Abstract

The chemical modification of L-2-haloacid halidohydrolase IVa (Hdl IVa), originally identified in Pseudomonas cepacia MBA4, produced as a recombinant protein in Escherichia coli DH5alpha, led to the identification of histidine and arginine as amino acid residues likely to play a part in the catalytic mechanism of the enzyme. These results, together with DNA sequence and analyses [Murdiyatmo, Asmara, Baines, Bull and Hardman (1992) Biochem. J. 284, 87-93] provided the basis for the rational design of a series of random- and site-directed-mutagenesis experiments of the Hdl IVa structural gene (hdl IVa). Subsequent apparent kinetic analyses of purified mutant enzymes identified His-20 and Arg-42 as the key residues in the activity of this halidohydrolase. It is also proposed that Asp-18 is implicated in the functioning of the enzyme, possibly by positioning the correct tautomer of His-20 for catalysis in the enzyme-substrate complex and stabilizing the protonated form of His-20 in the transition-state complex. Comparison of conserved amino acid sequences between the Hdl IVa and other halidohydrolases suggests that L-2-haloacid halidohydrolases contain conserved amino acid sequences that are not found in halidohydrolases active towards both D- and L-2-monochloropropionate.

Item Type: Article
Subjects: Q Science > QP Physiology (Living systems) > QP517 Biochemistry
Divisions: Faculties > Science Technology and Medical Studies > School of Biosciences
Depositing User: R.F. Xu
Date Deposited: 22 Oct 2009 22:43
Last Modified: 08 Jul 2014 10:19
Resource URI: https://kar.kent.ac.uk/id/eprint/20779 (The current URI for this page, for reference purposes)
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