5o9d: Difference between revisions

Line 1:

~~'''Unreleased structure'''~~

~~The entry~~ 5o9d is ~~ON HOLD until Paper~~ Publication

==Crystal structure of R. ruber ADH-A, mutant Y294F, W295A, Y54F, F43H, H39Y==

== Structural highlights ==

<table><tr><td colspan='2'>[[5o9d]] is a 4 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5O9D OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5O9D FirstGlance]. <br>

</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=9ON:(2~{S})-2-methylpentanedioic+acid'>9ON</scene>, <scene name='pdbligand=NAD:NICOTINAMIDE-ADENINE-DINUCLEOTIDE'>NAD</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr>

<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5o9d FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5o9d OCA], [http://pdbe.org/5o9d PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5o9d RCSB], [http://www.ebi.ac.uk/pdbsum/5o9d PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5o9d ProSAT]</span></td></tr>

</table>

== Publication Abstract from PubMed ==

Alcohol dehydrogenase A (ADH-A) from Rhodococcus ruber DSM 44541 is a promising biocatalyst for redox transformations of arylsubstituted sec-alcohols and ketones. The enzyme is stereoselective in the oxidation of 1-phenylethanol with a 300-fold preference for the (S)-enantiomer. The low catalytic efficiency with (R)-1-phenylethanol has been attributed to nonproductive binding of this substrate at the active site. Aiming to modify the enantioselectivity, to rather favor the (R)-alcohol, and also test the possible involvement of nonproductive substrate binding as a mechanism in substrate discrimination, we performed directed laboratory evolution of ADH-A. Three targeted sites that contribute to the active-site cavity were exposed to saturation mutagenesis in a stepwise manner and the generated variants were selected for improved catalytic activity with (R)-1-phenylethanol. After three subsequent rounds of mutagenesis, selection and structure-function analysis of isolated ADH-A variants, we conclude: (1) W295 has a key role as a structural determinant in the discrimination between (R)- and (S)-1-phenylethanol and a W295A substitution fundamentally changes the stereoselectivity of the protein. One observable effect is a faster rate of NADH release, which changes the rate-limiting step of the catalytic cycle from coenzyme release to hydride transfer. (2) The obtained change in enantiopreference, from the (S)- to the (R)-alcohol, can be partly explained by a shift in the nonproductive substrate binding modes. This article is protected by copyright. All rights reserved.

~~Authors: Dobritzsch~~, D., Maurer, D., ~~Hamnevik, E.~~, ~~Enugala~~, ~~T.R.~~, Widersten, M.

Relaxation of nonproductive binding and increased rate of coenzyme release in an alcohol dehydrogenase increases turnover with a non-preferred alcohol enantiomer.,Hamnevik E, Enugala TR, Maurer D, Ntuku S, Oliveira A, Dobritzsch D, Widersten M FEBS J. 2017 Sep 30. doi: 10.1111/febs.14279. PMID:28963762<ref>PMID:28963762</ref>

~~Description: Crystal structure~~ of R. ~~ruber ADH~~-~~A, mutant Y294F, W295A, Y54F, F43H~~, ~~H39Y~~

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br>

[[Category: ~~Unreleased Structures~~]]

</div>

== References ==

__TOC__

</StructureSection>

[[Category: Dobritzsch, D]]

[[Category: Enugala, T R]]

[[Category: Hamnevik, E]]

[[Category: Maurer, D]]

[[Category: Widersten, M]]

[[Category: ~~Enugala, T.R~~]]

[[Category: Alcohol dehydrogenase mutant variant]]

[[Category: ~~Maurer, D~~]]

[[Category: Nadh-dependent]]

[[Category: ~~Dobritzsch, D~~]]

[[Category: Oxidoreductase]]

[[Category: Rossmann fold]]

[[Category: Zn2+-dependent]]

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@@ Line 1: / Line 1: @@
-'''Unreleased structure'''
-The entry 5o9d is ON HOLD  until Paper Publication
+==Crystal structure of R. ruber ADH-A, mutant Y294F, W295A, Y54F, F43H, H39Y==
+<StructureSection load='5o9d' size='340' side='right' caption='[[5o9d]], [[Resolution|resolution]] 1.79&Aring;' scene=''>
+== Structural highlights ==
+<table><tr><td colspan='2'>[[5o9d]] is a 4 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5O9D OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5O9D FirstGlance]. <br>
+</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=9ON:(2~{S})-2-methylpentanedioic+acid'>9ON</scene>, <scene name='pdbligand=NAD:NICOTINAMIDE-ADENINE-DINUCLEOTIDE'>NAD</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr>
+<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5o9d FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5o9d OCA], [http://pdbe.org/5o9d PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5o9d RCSB], [http://www.ebi.ac.uk/pdbsum/5o9d PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5o9d ProSAT]</span></td></tr>
+</table>
+<div style="background-color:#fffaf0;">
+== Publication Abstract from PubMed ==
+Alcohol dehydrogenase A (ADH-A) from Rhodococcus ruber DSM 44541 is a promising biocatalyst for redox transformations of arylsubstituted sec-alcohols and ketones. The enzyme is stereoselective in the oxidation of 1-phenylethanol with a 300-fold preference for the (S)-enantiomer. The low catalytic efficiency with (R)-1-phenylethanol has been attributed to nonproductive binding of this substrate at the active site. Aiming to modify the enantioselectivity, to rather favor the (R)-alcohol, and also test the possible involvement of nonproductive substrate binding as a mechanism in substrate discrimination, we performed directed laboratory evolution of ADH-A. Three targeted sites that contribute to the active-site cavity were exposed to saturation mutagenesis in a stepwise manner and the generated variants were selected for improved catalytic activity with (R)-1-phenylethanol. After three subsequent rounds of mutagenesis, selection and structure-function analysis of isolated ADH-A variants, we conclude: (1) W295 has a key role as a structural determinant in the discrimination between (R)- and (S)-1-phenylethanol and a W295A substitution fundamentally changes the stereoselectivity of the protein. One observable effect is a faster rate of NADH release, which changes the rate-limiting step of the catalytic cycle from coenzyme release to hydride transfer. (2) The obtained change in enantiopreference, from the (S)- to the (R)-alcohol, can be partly explained by a shift in the nonproductive substrate binding modes. This article is protected by copyright. All rights reserved.
-Authors: Dobritzsch, D., Maurer, D., Hamnevik, E., Enugala, T.R., Widersten, M.
+Relaxation of nonproductive binding and increased rate of coenzyme release in an alcohol dehydrogenase increases turnover with a non-preferred alcohol enantiomer.,Hamnevik E, Enugala TR, Maurer D, Ntuku S, Oliveira A, Dobritzsch D, Widersten M FEBS J. 2017 Sep 30. doi: 10.1111/febs.14279. PMID:28963762<ref>PMID:28963762</ref>
-Description: Crystal structure of R. ruber ADH-A, mutant Y294F, W295A, Y54F, F43H, H39Y
+From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
-[[Category: Unreleased Structures]]
+</div>
+<div class="pdbe-citations 5o9d" style="background-color:#fffaf0;"></div>
+== References ==
+<references/>
+__TOC__
+</StructureSection>
+[[Category: Dobritzsch, D]]
+[[Category: Enugala, T R]]
 [[Category: Hamnevik, E]]
+[[Category: Maurer, D]]
 [[Category: Widersten, M]]
-[[Category: Enugala, T.R]]
+[[Category: Alcohol dehydrogenase mutant variant]]
-[[Category: Maurer, D]]
+[[Category: Nadh-dependent]]
-[[Category: Dobritzsch, D]]
+[[Category: Oxidoreductase]]
+[[Category: Rossmann fold]]
+[[Category: Zn2+-dependent]]