1e0y
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(New page: 200px<br /><applet load="1e0y" size="450" color="white" frame="true" align="right" spinBox="true" caption="1e0y, resolution 2.75Å" /> '''STRUCTURE OF THE D17...)
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Revision as of 11:36, 20 November 2007
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STRUCTURE OF THE D170S/T457E DOUBLE MUTANT OF VANILLYL-ALCOHOL OXIDASE
Overview
Vanillyl-alcohol oxidase (VAO) is the prototype of a newly recognized, family of structurally related oxidoreductases sharing a conserved, FAD-binding domain. The active site of VAO is formed by a cavity where the, enzyme is able to catalyze many reactions with phenolic substrates. Among, these reactions is the stereospecific hydroxylation of, 4-ethylphenol-forming (R)-1-(4'-hydroxyphenyl)ethanol. During this, conversion, Asp-170 is probably critical for the hydration of the, initially formed p-quinone methide intermediate. By site-directed, mutagenesis, the putative active site base has been relocated to the, opposite face of the active site cavity. In this way, a change in, stereospecificity has been achieved. Like native VAO, the single mutants, T457E, D170A, and D170S preferentially converted 4-ethylphenol to the, (R)-enantiomer of 1-(4'-hydroxyphenyl)ethanol. The double mutants, D170A/T457E and D170S/T457E exhibited an inverted stereospecificity with, 4-ethylphenol. Particularly, D170S/T457E was strongly (S)-selective, with, an enantiomeric excess of 80%. The crystal structure of D170S/T457E, in, complex with trifluoromethylphenol, showed a highly conserved mode of, ligand binding and revealed that the distinctive catalytic properties of, this mutant are not caused by major structural changes.
About this Structure
1E0Y is a Single protein structure of sequence from Penicillium simplicissimum with FAD and FCR as ligands. Active as Vanillyl-alcohol oxidase, with EC number 1.1.3.38 Full crystallographic information is available from OCA.
Reference
Inversion of stereospecificity of vanillyl-alcohol oxidase., van Den Heuvel RH, Fraaije MW, Ferrer M, Mattevi A, van Berkel WJ, Proc Natl Acad Sci U S A. 2000 Aug 15;97(17):9455-60. PMID:10920192
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