View source for 8grj


==Crystal structure of gamma-alpha subunit complex from Burkholderia cepacia FAD glucose dehydrogenase in complex with gluconolactone==
<StructureSection load='8grj' size='340' side='right'caption='[[8grj]], [[Resolution|resolution]] 2.95&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[8grj]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Burkholderia_cepacia Burkholderia cepacia]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=8GRJ OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=8GRJ 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]] 2.95&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>, <scene name='pdbligand=LGC:(3S,4R,5R,6S)-3,4,5-TRIHYDROXY-6-(HYDROXYMETHYL)TETRAHYDRO-2H-PYRAN-2-ONE'>LGC</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=8grj FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=8grj OCA], [https://pdbe.org/8grj PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=8grj RCSB], [https://www.ebi.ac.uk/pdbsum/8grj PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=8grj ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/Q8GQE7_BURCE Q8GQE7_BURCE] 
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
The heterotrimeric flavin adenine dinucleotide (FAD) dependent glucose dehydrogenase derived from Burkholderia cepacia (BcGDH) has many exceptional features for its use in glucose sensing-including that this enzyme is capable of direct electron transfer with an electrode in its heterotrimeric configuration. However, this enzyme's high catalytic activity towards not only glucose but also galactose presents an engineering challenge. To increase the substrate specificity of this enzyme, it must be engineered to reduce its activity towards galactose while maintaining its activity towards glucose. To aid in these mutagenesis studies, the crystal structure composed of BcGDH's small subunit and catalytic subunit (BcGDHgammaalpha), in complex with D-glucono-1,5-lactone was elucidated and used to construct the three-dimensional model for targeted, site-directed mutagenesis. BcGDHgammaalpha was then mutated at three different residues, glycine 322, asparagine 474 and asparagine 475. The single mutations that showed the greatest glucose selectivity were combined to create the resulting mutant, alpha-G322Q-N474S-N475S. The alpha-G322Q-N474S-N475S mutant and BcGDHgammaalpha wild type were then characterized with dye-mediated dehydrogenase activity assays to determine their kinetic parameters. The alpha-G322Q-N474S-N475S mutant showed more than a 2-fold increase in V(max) towards glucose and this mutant showed a lower activity towards galactose in the physiological range (5â¯mM) of 4.19â¯Uâ¯mg(-1), as compared to the wild type, 86.6â¯Uâ¯mg(-1). This resulting increase in specificity lead to an 81.7â¯gal/glc % activity for the wild type while the alpha-G322Q-N474S-N475S mutant had just 10.9â¯gal/glc % activity at 5â¯mM. While the BcGDHgammaalpha wild type has high specificity towards galactose, our engineering alpha-G322Q-N474S-N475S mutant showed concentration dependent response to glucose and was not affected by galactose.

Improvement of substrate specificity of the direct electron transfer type FAD-dependent glucose dehydrogenase catalytic subunit.,Kerrigan JA Jr, Yoshida H, Okuda-Shimazaki J, Temple B, Kojima K, Sode K J Biotechnol. 2024 Sep 25;395:170-179. doi: 10.1016/j.jbiotec.2024.09.013. PMID:39326560<ref>PMID:39326560</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 8grj" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Burkholderia cepacia]]
[[Category: Large Structures]]
[[Category: Kerrigan JA]]
[[Category: Kojima K]]
[[Category: Okuda-Shimazaki J]]
[[Category: Sode K]]
[[Category: Tsugawa W]]
[[Category: Yoshida H]]