7zzk
From Proteopedia
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== Structural highlights == | == Structural highlights == | ||
<table><tr><td colspan='2'>[[7zzk]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Ralstonia_solanacearum Ralstonia solanacearum]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=7ZZK OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=7ZZK FirstGlance]. <br> | <table><tr><td colspan='2'>[[7zzk]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Ralstonia_solanacearum Ralstonia solanacearum]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=7ZZK OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=7ZZK FirstGlance]. <br> | ||
| - | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ALA:ALANINE'>ALA</scene>, <scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>, <scene name='pdbligand=GOL:GLYCEROL'>GOL</scene>, <scene name='pdbligand=PEG:DI(HYDROXYETHYL)ETHER'>PEG</scene></td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.5Å</td></tr> |
| + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ALA:ALANINE'>ALA</scene>, <scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>, <scene name='pdbligand=GOL:GLYCEROL'>GOL</scene>, <scene name='pdbligand=PEG:DI(HYDROXYETHYL)ETHER'>PEG</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=7zzk FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=7zzk OCA], [https://pdbe.org/7zzk PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=7zzk RCSB], [https://www.ebi.ac.uk/pdbsum/7zzk PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=7zzk ProSAT]</span></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=7zzk FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=7zzk OCA], [https://pdbe.org/7zzk PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=7zzk RCSB], [https://www.ebi.ac.uk/pdbsum/7zzk PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=7zzk ProSAT]</span></td></tr> | ||
</table> | </table> | ||
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Flavin-dependent carbohydrate oxidases are valuable tools in biotechnological applications due to their high selectivity in the oxidation of carbohydrates. In this study, we report the biochemical and structural characterization of a recently discovered carbohydrate oxidase from the bacterium Ralstonia solanacearum, which is a member of the vanillyl alcohol oxidase flavoprotein family. Due to its exceptionally high activity toward N-acetyl-d-galactosamine and N-acetyl-d-glucosamine, the enzyme was named N-acetyl-glucosamine oxidase (NagOx). In contrast to most known (fungal) carbohydrate oxidases, NagOx could be overexpressed in a bacterial host, which facilitated detailed biochemical and enzyme engineering studies. Steady state kinetic analyses revealed that non-acetylated hexoses were also accepted as substrates albeit with lower efficiency. Upon determination of the crystal structure, structural insights into NagOx were obtained. A large cavity containing a bicovalently bound FAD, tethered via histidyl and cysteinyl linkages, was observed. Substrate docking highlighted how a single residue (Leu251) plays a key role in the accommodation of N-acetylated sugars in the active site. Upon replacement of Leu251 (L251R mutant), an enzyme variant was generated with a drastically modified substrate acceptance profile, tuned toward non-N-acetylated monosaccharides and disaccharides. Furthermore, the activity toward bulkier substrates such as the trisaccharide maltotriose was introduced by this mutation. Due to its advantage of being overexpressed in a bacterial host, NagOx can be considered a promising alternative engineerable biocatalyst for selective oxidation of monosaccharides and oligosaccharides. | Flavin-dependent carbohydrate oxidases are valuable tools in biotechnological applications due to their high selectivity in the oxidation of carbohydrates. In this study, we report the biochemical and structural characterization of a recently discovered carbohydrate oxidase from the bacterium Ralstonia solanacearum, which is a member of the vanillyl alcohol oxidase flavoprotein family. Due to its exceptionally high activity toward N-acetyl-d-galactosamine and N-acetyl-d-glucosamine, the enzyme was named N-acetyl-glucosamine oxidase (NagOx). In contrast to most known (fungal) carbohydrate oxidases, NagOx could be overexpressed in a bacterial host, which facilitated detailed biochemical and enzyme engineering studies. Steady state kinetic analyses revealed that non-acetylated hexoses were also accepted as substrates albeit with lower efficiency. Upon determination of the crystal structure, structural insights into NagOx were obtained. A large cavity containing a bicovalently bound FAD, tethered via histidyl and cysteinyl linkages, was observed. Substrate docking highlighted how a single residue (Leu251) plays a key role in the accommodation of N-acetylated sugars in the active site. Upon replacement of Leu251 (L251R mutant), an enzyme variant was generated with a drastically modified substrate acceptance profile, tuned toward non-N-acetylated monosaccharides and disaccharides. Furthermore, the activity toward bulkier substrates such as the trisaccharide maltotriose was introduced by this mutation. Due to its advantage of being overexpressed in a bacterial host, NagOx can be considered a promising alternative engineerable biocatalyst for selective oxidation of monosaccharides and oligosaccharides. | ||
| - | Structural Elucidation and Engineering of a Bacterial Carbohydrate Oxidase.,Boverio A, Widodo WS, Santema LL, Rozeboom HJ, Xiang R, Guallar V, Mattevi A, Fraaije MW Biochemistry. | + | Structural Elucidation and Engineering of a Bacterial Carbohydrate Oxidase.,Boverio A, Widodo WS, Santema LL, Rozeboom HJ, Xiang R, Guallar V, Mattevi A, Fraaije MW Biochemistry. 2023 Jan 17;62(2):429-436. doi: 10.1021/acs.biochem.2c00307. Epub , 2022 Jul 26. PMID:35881507<ref>PMID:35881507</ref> |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | ||
Current revision
Structure of the N-acetyl-D-glucosamine oxidase from Ralstonia Solanacearum
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