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| - | [[Image:1w1m.gif|left|200px]] | + | {{Seed}} |
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| | {{STRUCTURE_1w1m| PDB=1w1m | SCENE= }} | | {{STRUCTURE_1w1m| PDB=1w1m | SCENE= }} |
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| - | '''STRUCTURE OF THE OCTAMERIC FLAVOENZYME VANILLYL-ALCOHOL OXIDASE: GLU502GLY MUTANT'''
| + | ===STRUCTURE OF THE OCTAMERIC FLAVOENZYME VANILLYL-ALCOHOL OXIDASE: GLU502GLY MUTANT=== |
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| - | ==Overview==
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| - | The flavoenzyme vanillyl-alcohol oxidase was subjected to random mutagenesis to generate mutants with enhanced reactivity to creosol (2-methoxy-4-methylphenol). The vanillyl-alcohol oxidase-mediated conversion of creosol proceeds via a two-step process in which the initially formed vanillyl alcohol (4-hydroxy-3-methoxybenzyl alcohol) is oxidized to the widely used flavor compound vanillin (4-hydroxy-3-methoxybenzaldehyde). The first step of this reaction is extremely slow due to the formation of a covalent FAD N-5-creosol adduct. After a single round of error-prone PCR, seven mutants were generated with increased reactivity to creosol. The single-point mutants I238T, F454Y, E502G, and T505S showed an up to 40-fold increase in catalytic efficiency (kcat/Km) with creosol compared with the wild-type enzyme. This enhanced reactivity was due to a lower stability of the covalent flavin-substrate adduct, thereby promoting vanillin formation. The catalytic efficiencies of the mutants were also enhanced for other ortho-substituted 4-methylphenols, but not for p-cresol (4-methylphenol). The replaced amino acid residues are not located within a distance of direct interaction with the substrate, and the determined three-dimensional structures of the mutant enzymes are highly similar to that of the wild-type enzyme. These results clearly show the importance of remote residues, not readily predicted by rational design, for the substrate specificity of enzymes.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_15169773}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 15169773 is the PubMed ID number. |
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| | + | {{ABSTRACT_PUBMED_15169773}} |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Flavoenzyme]] | | [[Category: Flavoenzyme]] |
| | [[Category: Oxidoreductase]] | | [[Category: Oxidoreductase]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May 3 13:01:40 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Jul 28 15:30:30 2008'' |
Revision as of 12:30, 28 July 2008
Template:STRUCTURE 1w1m
STRUCTURE OF THE OCTAMERIC FLAVOENZYME VANILLYL-ALCOHOL OXIDASE: GLU502GLY MUTANT
Template:ABSTRACT PUBMED 15169773
About this Structure
1W1M is a Single protein structure of sequence from Penicillium simplicissimum. Full crystallographic information is available from OCA.
Reference
Laboratory-evolved vanillyl-alcohol oxidase produces natural vanillin., van den Heuvel RH, van den Berg WA, Rovida S, van Berkel WJ, J Biol Chem. 2004 Aug 6;279(32):33492-500. Epub 2004 May 28. PMID:15169773
Page seeded by OCA on Mon Jul 28 15:30:30 2008
