1w1j: Difference between revisions

Revision as of 13:01, 3 May 2008

STRUCTURE OF THE OCTAMERIC FLAVOENZYME VANILLYL-ALCOHOL OXIDASE: THE505SER MUTANT

OverviewOverview

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.

About this StructureAbout this Structure

1W1J is a Single protein structure of sequence from Penicillium simplicissimum. Full crystallographic information is available from OCA.

ReferenceReference

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 Sat May 3 13:01:37 2008

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OCA

@@ Line 1: / Line 1: @@
 [[Image:1w1j.gif|left|200px]]
- {{Structure
+<!--
-|PDB= 1w1j |SIZE=350|CAPTION= <scene name='initialview01'>1w1j</scene>, resolution 2.70&Aring;
+The line below this paragraph, containing "STRUCTURE_1w1j", creates the "Structure Box" on the page.
-|SITE= <scene name='pdbsite=AC1:Eug+Binding+Site+For+Chain+B'>AC1</scene>
+You may change the PDB parameter (which sets the PDB file loaded into the applet)
-|LIGAND= <scene name='pdbligand=EUG:2-METHOXY-4-VINYL-PHENOL'>EUG</scene>, <scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>
+or the SCENE parameter (which sets the initial scene displayed when the page is loaded),
-|ACTIVITY= <span class='plainlinks'>[http://en.wikipedia.org/wiki/Alcohol_oxidase Alcohol oxidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.1.3.13 1.1.3.13] </span>
+or leave the SCENE parameter empty for the default display.
-|GENE=
+-->
-|DOMAIN=
+{{STRUCTURE_1w1j|  PDB=1w1j  |  SCENE=  }}
-|RELATEDENTRY=
-|RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1w1j FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1w1j OCA], [http://www.ebi.ac.uk/pdbsum/1w1j PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=1w1j RCSB]</span>
-}}
 '''STRUCTURE OF THE OCTAMERIC FLAVOENZYME VANILLYL-ALCOHOL OXIDASE: THE505SER MUTANT'''
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 [[Category: Single protein]]
 [[Category: Heuvel, R H.Van Den.]]
-[[Category: catalysis]]
+[[Category: Catalysis]]
-[[Category: fad]]
+[[Category: Fad]]
-[[Category: flavoenzyme]]
+[[Category: Flavoenzyme]]
-[[Category: oxidoreductase]]
+[[Category: Oxidoreductase]]
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May  3 13:01:37 2008''
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Mar 31 00:29:48 2008''