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| [[Image:1z9a.gif|left|200px]]<br /><applet load="1z9a" size="350" color="white" frame="true" align="right" spinBox="true"
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| caption="1z9a, resolution 2.40Å" />
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| '''Crystal Structure Of The Asn-309 To Asp Mutant Of Candida Tenuis Xylose Reductase (Akr2B5) Bound To Nad+'''<br />
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| ==Overview== | | ==Crystal Structure Of The Asn-309 To Asp Mutant Of Candida Tenuis Xylose Reductase (Akr2B5) Bound To Nad+== |
| Little is known about how substrates bind to CtXR (Candida tenuis xylose reductase; AKR2B5) and other members of the AKR (aldo-keto reductase) protein superfamily. Modelling of xylose into the active site of CtXR suggested that Trp23, Asp50 and Asn309 are the main components of pentose-specific substrate-binding recognition. Kinetic consequences of site-directed substitutions of these residues are reported. The mutants W23F and W23Y catalysed NADH-dependent reduction of xylose with only 4 and 1% of the wild-type efficiency (kcat/K(m)) respectively, but improved the wild-type selectivity for utilization of ketones, relative to xylose, by factors of 156 and 471 respectively. Comparison of multiple sequence alignment with reported specificities of AKR members emphasizes a conserved role of Trp23 in determining aldehyde-versus-ketone substrate selectivity. D50A showed 31 and 18% of the wild-type catalytic-centre activities for xylose reduction and xylitol oxidation respectively, consistent with a decrease in the rates of the chemical steps caused by the mutation, but no change in the apparent substrate binding constants and the pattern of substrate specificities. The 30-fold preference of the wild-type for D-galactose compared with 2-deoxy-D-galactose was lost completely in N309A and N309D mutants. Comparison of the 2.4 A (1 A=0.1 nm) X-ray crystal structure of mutant N309D bound to NAD+ with the previous structure of the wild-type holoenzyme reveals no major structural perturbations. The results suggest that replacement of Asn309 with alanine or aspartic acid disrupts the function of the original side chain in donating a hydrogen atom for bonding with the substrate C-2(R) hydroxy group, thus causing a loss of transition-state stabilization energy of 8-9 kJ/mol.
| | <StructureSection load='1z9a' size='340' side='right'caption='[[1z9a]], [[Resolution|resolution]] 2.40Å' scene=''> |
| | | == Structural highlights == |
| ==About this Structure== | | <table><tr><td colspan='2'>[[1z9a]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Yamadazyma_tenuis Yamadazyma tenuis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1Z9A OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1Z9A FirstGlance]. <br> |
| 1Z9A is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Candida_tenuis Candida tenuis] with <scene name='pdbligand=NAD:'>NAD</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1Z9A OCA].
| | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.4Å</td></tr> |
| | | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=NAD:NICOTINAMIDE-ADENINE-DINUCLEOTIDE'>NAD</scene></td></tr> |
| ==Reference==
| | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1z9a FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1z9a OCA], [https://pdbe.org/1z9a PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1z9a RCSB], [https://www.ebi.ac.uk/pdbsum/1z9a PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1z9a ProSAT]</span></td></tr> |
| Probing the substrate binding site of Candida tenuis xylose reductase (AKR2B5) with site-directed mutagenesis., Kratzer R, Leitgeb S, Wilson DK, Nidetzky B, Biochem J. 2006 Jan 1;393(Pt 1):51-8. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=16336198 16336198]
| | </table> |
| [[Category: Candida tenuis]] | | == Function == |
| [[Category: Single protein]]
| | [https://www.uniprot.org/uniprot/XYL1_CANTE XYL1_CANTE] Reduces D-xylose into xylitol. Has a preference for NADPH, but can also utilize NADH as cosubstrate. |
| [[Category: Kratzer, R.]] | | == Evolutionary Conservation == |
| [[Category: Leitgeb, S.]]
| | [[Image:Consurf_key_small.gif|200px|right]] |
| [[Category: Nidetzky, B.]]
| | Check<jmol> |
| [[Category: Wilson, D K.]] | | <jmolCheckbox> |
| [[Category: NAD]] | | <scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/z9/1z9a_consurf.spt"</scriptWhenChecked> |
| [[Category: akr]] | | <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked> |
| [[Category: aldo-keto reductase]]
| | <text>to colour the structure by Evolutionary Conservation</text> |
| [[Category: beta-alpha-barrel]] | | </jmolCheckbox> |
| [[Category: candida tenuis]] | | </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/main_output.php?pdb_ID=1z9a ConSurf]. |
| [[Category: ketone reduction]] | | <div style="clear:both"></div> |
| [[Category: structure-activity correlation]] | | __TOC__ |
| [[Category: substrate selectivity]] | | </StructureSection> |
| [[Category: xylose reductase]] | | [[Category: Large Structures]] |
| | | [[Category: Yamadazyma tenuis]] |
| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 16:13:19 2008''
| | [[Category: Kratzer R]] |
| | [[Category: Leitgeb S]] |
| | [[Category: Nidetzky B]] |
| | [[Category: Wilson DK]] |
