1r38: Difference between revisions
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[[Image: | ==Crystal structure of H114A mutant of Candida tenuis xylose reductase== | ||
<StructureSection load='1r38' size='340' side='right' caption='[[1r38]], [[Resolution|resolution]] 2.20Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[1r38]] is a 4 chain structure with sequence from [http://en.wikipedia.org/wiki/Candida_tenuis Candida tenuis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1R38 OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1R38 FirstGlance]. <br> | |||
</td></tr><tr><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=NAP:NADP+NICOTINAMIDE-ADENINE-DINUCLEOTIDE+PHOSPHATE'>NAP</scene><br> | |||
<tr><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[1jez|1jez]], [[1k8c|1k8c]], [[1mi3|1mi3]]</td></tr> | |||
<tr><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">xylr ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=45596 Candida tenuis])</td></tr> | |||
<tr><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Aldehyde_reductase Aldehyde reductase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.1.1.21 1.1.1.21] </span></td></tr> | |||
<tr><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1r38 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1r38 OCA], [http://www.rcsb.org/pdb/explore.do?structureId=1r38 RCSB], [http://www.ebi.ac.uk/pdbsum/1r38 PDBsum]</span></td></tr> | |||
<table> | |||
== Evolutionary Conservation == | |||
[[Image:Consurf_key_small.gif|200px|right]] | |||
Check<jmol> | |||
<jmolCheckbox> | |||
<scriptWhenChecked>select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/r3/1r38_consurf.spt"</scriptWhenChecked> | |||
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked> | |||
<text>to colour the structure by Evolutionary Conservation</text> | |||
</jmolCheckbox> | |||
</jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/chain_selection.php?pdb_ID=2ata ConSurf]. | |||
<div style="clear:both"></div> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Xylose reductase from the yeast Candida tenuis (CtXR) is a family 2 member of the aldo-keto reductase (AKR) superfamily of proteins and enzymes. Active site His-113 is conserved among AKRs, but a unified mechanism of how it affects catalytic activity is outstanding. We have replaced His-113 by alanine using site-directed mutagenesis, determined a 2.2 A structure of H113A mutant bound to NADP(+), and compared catalytic reaction profiles of NADH-dependent reduction of different aldehydes catalyzed by the wild type and the mutant. Deuterium kinetic isotope effects (KIEs) on k(cat) and k(cat)/K(m xylose) show that, relative to the wild type, the hydride transfer rate constant (k(7) approximately 0.16 s(-1)) has decreased about 1000-fold in H113A whereas xylose binding was not strongly affected. No solvent isotope effect was seen on k(cat) and k(cat)/K(m xylose) for H113A, suggesting that proton transfer has not become rate-limiting as a result of the mutation. The pH profiles of log(k(cat)/K(m xylose)) for the wild type and H113A decreased above apparent pK(a) values of 8.85 and 7.63, respectively. The DeltapK(a) of -1.2 pH units likely reflects a proximally disruptive character of the mutation, affecting the position of Asp-50. A steady-state kinetic analysis for H113A-catalyzed reduction of a homologous series of meta-substituted benzaldehyde derivatives was carried out, and quantitative structure-reactivity correlations were used to factor the observed kinetic substituent effect on k(cat) and k(cat)/K(m aldehyde) into an electronic effect and bonding effects (which are lacking in the wild type). Using the Hammett sigma scale, electronic parameter coefficients (rho) of +0.64 (k(cat)) and +0.78 (k(cat)/K(m aldehyde)) were calculated and clearly differ from rho(k(cat)/K(aldehyde)) and rho(k(cat)) values of +1.67 and approximately 0.0, respectively, for the wild-type enzyme. Hydride transfer rate constants of H113A, calculated from kinetic parameters and KIE data, display a substituent dependence not seen in the corresponding wild-type enzyme rate constants. An enzymic mechanism is proposed in which His-113, through a hydrogen bond from Nepsilon2 to aldehyde O1, assists in catalysis by optimizing the C=O bond charge separation and orbital alignment in the ternary complex. | |||
Studies of the enzymic mechanism of Candida tenuis xylose reductase (AKR 2B5): X-ray structure and catalytic reaction profile for the H113A mutant.,Kratzer R, Kavanagh KL, Wilson DK, Nidetzky B Biochemistry. 2004 May 4;43(17):4944-54. PMID:15109252<ref>PMID:15109252</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
== | |||
< | |||
[[Category: Aldehyde reductase]] | [[Category: Aldehyde reductase]] | ||
[[Category: Candida tenuis]] | [[Category: Candida tenuis]] | ||
