4rnx: Difference between revisions
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==K154 Circular Permutation of Old Yellow Enzyme== | |||
<StructureSection load='4rnx' size='340' side='right'caption='[[4rnx]], [[Resolution|resolution]] 1.25Å' scene=''> | |||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[4rnx]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Saccharomyces_pastorianus Saccharomyces pastorianus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4RNX OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4RNX FirstGlance]. <br> | |||
</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.25Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=EDO:1,2-ETHANEDIOL'>EDO</scene>, <scene name='pdbligand=FMN:FLAVIN+MONONUCLEOTIDE'>FMN</scene></td></tr> | |||
<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=4rnx FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4rnx OCA], [https://pdbe.org/4rnx PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4rnx RCSB], [https://www.ebi.ac.uk/pdbsum/4rnx PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4rnx ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/OYE1_SACPS OYE1_SACPS] Oxidizes beta-NADH, beta-NADPH, and alpha-NADPH. | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Circular permutation of the NADPH-dependent oxidoreductase Old Yellow Enzyme from Saccharomyces pastorianus (OYE1) can significantly enhance the enzyme's catalytic performance. Termini relocation into four regions of the protein (sectors I-IV) near the active site has proven effective in altering enzyme function. To better understand the structural consequences and rationalize the observed functional gains in these OYE1 variants, we selected representatives from sectors I-III for further characterization by biophysical methods and X-ray crystallography. These investigations not only show trends in enzyme stability and quaternary structure as a function of termini location, but also provide a possible explanation for the catalytic gains in our top-performing OYE variant (new N-terminus at residue 303; sector III). Crystallographic analysis indicates that termini relocation into sector III affects the loop beta6 region (amino acid positions: 290-310) of OYE1 which forms a lid over the active site. Peptide backbone cleavage greatly enhances local flexibility, effectively converting the loop into a tether and consequently increasing the environmental exposure of the active site. Interestingly, such active site remodeling does not negatively impact the enzyme's activity and stereoselectivity, nor does it perturb the conformation of other key active site residues with the exception of Y375. These observations were confirmed in truncation experiments, deleting all residues of the loop beta6 region in our OYE variant. Intrigued by the finding that circular permutation leaves most of the key catalytic residues unchanged, we also tested OYE permutants for possible additive or synergistic effects of amino acid substitutions. Distinct functional changes in these OYE variants were detected upon mutations at W116, known in native OYE1 to cause inversion of diastereo-selectivity for (S)-carvone reduction. Our findings demonstrate the contribution of loop beta6 toward determining the stereoselectivity of OYE1, an important insight for future OYE engineering efforts. | |||
STRUCTURAL AND FUNCTIONAL CONSEQUENCES OF CIRCULAR PERMUTATION ON THE ACTIVE SITE OF OLD YELLOW ENZYME.,Daugherty AB, Horton JR, Cheng X, Lutz S ACS Catal. 2015 Feb 6;5(2):892-899. doi: 10.1021/cs501702k. PMID:25692074<ref>PMID:25692074</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
[[Category: | </div> | ||
[[Category: | <div class="pdbe-citations 4rnx" style="background-color:#fffaf0;"></div> | ||
[[Category: | |||
[[Category: | ==See Also== | ||
[[Category: Horton | *[[NADPH dehydrogenase|NADPH dehydrogenase]] | ||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Saccharomyces pastorianus]] | |||
[[Category: Cheng X]] | |||
[[Category: Daugherty AB]] | |||
[[Category: Horton JR]] | |||
[[Category: Lutz S]] | |||