1t2x
From Proteopedia
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Glactose oxidase C383S mutant identified by directed evolution
Overview
Galactose oxidase (GO; E.C. 1.1.3.9) is a copper- containing enzyme that, oxidizes a range of primary alcohols to aldehydes. This broad substrate, specificity is reflected in a high K(M) for substrates. Directed evolution, has previously been used to select variants of GO that exhibit enhanced, expression and kinetic properties. In assays using unpurified enzyme, samples, the variant C383S displayed a 5-fold lower K(M) than wild-type, GO. In the present study, we have constructed, expressed, purified and, characterized a number of single, double and triple mutants at residues, Cys383, Tyr436 and Val494, identified in one of the directed evolution, studies, to examine their relative contributions to improved catalytic, activity of GO. We report kinetic studies on the various mutant enzymes., In addition, we have determined the three-dimensional structure of the, C383S variant. As with many mutations identified in directed evolution, experiments, the availability of structural information does not provide a, definitive answer to the reason for the improved K(M) in the C383S variant, protein.
About this Structure
1T2X is a Protein complex structure of sequences from Fusarium sp. with NA, CU and ACT as ligands. Active as Galactose oxidase, with EC number 1.1.3.9 Full crystallographic information is available from OCA.
Reference
Structural and kinetic studies of a series of mutants of galactose oxidase identified by directed evolution., Wilkinson D, Akumanyi N, Hurtado-Guerrero R, Dawkes H, Knowles PF, Phillips SE, McPherson MJ, Protein Eng Des Sel. 2004 Feb;17(2):141-8. Epub 2004 Jan 12. PMID:15047910
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