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==Crystal structure of rat xanthine oxidoreductase mutant (W335A and F336L)== | |||
<StructureSection load='2e3t' size='340' side='right'caption='[[2e3t]], [[Resolution|resolution]] 2.28Å' scene=''> | |||
| | == Structural highlights == | ||
<table><tr><td colspan='2'>[[2e3t]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Rattus_norvegicus Rattus norvegicus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=2E3T OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=2E3T 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]] 2.28Å</td></tr> | |||
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=BCT:BICARBONATE+ION'>BCT</scene>, <scene name='pdbligand=CA:CALCIUM+ION'>CA</scene>, <scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene>, <scene name='pdbligand=FES:FE2/S2+(INORGANIC)+CLUSTER'>FES</scene>, <scene name='pdbligand=PO4:PHOSPHATE+ION'>PO4</scene>, <scene name='pdbligand=URC:URIC+ACID'>URC</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=2e3t FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=2e3t OCA], [https://pdbe.org/2e3t PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=2e3t RCSB], [https://www.ebi.ac.uk/pdbsum/2e3t PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=2e3t ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/XDH_RAT XDH_RAT] Key enzyme in purine degradation. Catalyzes the oxidation of hypoxanthine to xanthine. Catalyzes the oxidation of xanthine to uric acid. Contributes to the generation of reactive oxygen species.<ref>PMID:17301076</ref> | |||
== Evolutionary Conservation == | |||
[[Image:Consurf_key_small.gif|200px|right]] | |||
Check<jmol> | |||
<jmolCheckbox> | |||
<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/e3/2e3t_consurf.spt"</scriptWhenChecked> | |||
<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked> | |||
<text>to colour the structure by Evolutionary Conservation</text> | |||
</jmolCheckbox> | |||
</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=2e3t ConSurf]. | |||
<div style="clear:both"></div> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
Reactive oxygen species are generated by various systems, including NADPH oxidases, xanthine oxidoreductase (XOR) and mitochondrial respiratory enzymes, and contribute to many physiological and pathological phenomena. Mammalian xanthine dehydrogenase (XDH) can be converted to xanthine oxidase (XO), which produces both superoxide anion and hydrogen peroxide in a molar ratio of about 1:3, depending upon the conditions. Here, we present a mutant of rat XOR that displays mainly XO activity with a superoxide:hydrogen peroxide production ratio of about 6:1. In the mutant, tryptophan 335, which is a component of the amino acid cluster crucial for switching from the XDH to the XO conformation, was replaced with alanine, and phenylalanine 336, which modulates FAD's redox potential through stacking interactions with the flavin cofactor, was changed to leucine. When the mutant was expressed in Sf9 cells, it was obtained in the XO form, and dithiothreitol treatment only partially restored the pyridine nucleotide-binding capacity. The crystal structure of the dithiothreitol-treated mutant at 2.3 Angstroms resolution showed the enzyme's two subunits to be quite similar, but not identical: the cluster involved in conformation-switching was completely disrupted in one subunit, but remained partly associated in the other one. The chain trace of the active site loop in this mutant is very similar to that of the bovine XO form. These results are consistent with the idea that the XDH and XO forms of the mutant are in an equilibrium that greatly favours the XO form, but the equilibrium is partly shifted towards the XDH form upon incubation with dithiothreitol. | |||
Two mutations convert mammalian xanthine oxidoreductase to highly superoxide-productive xanthine oxidase.,Asai R, Nishino T, Matsumura T, Okamoto K, Igarashi K, Pai EF, Nishino T J Biochem. 2007 Apr;141(4):525-34. Epub 2007 Feb 14. PMID:17301076<ref>PMID:17301076</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 2e3t" style="background-color:#fffaf0;"></div> | |||
== | ==See Also== | ||
*[[Xanthine dehydrogenase 3D structures|Xanthine dehydrogenase 3D structures]] | |||
== References == | |||
== | <references/> | ||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Rattus norvegicus]] | [[Category: Rattus norvegicus]] | ||
[[Category: Asai R]] | |||
[[Category: Asai | [[Category: Matsumura T]] | ||
[[Category: Matsumura | [[Category: Nishino T]] | ||
[[Category: Nishino | [[Category: Okamoto K]] | ||
[[Category: Okamoto | [[Category: Pai EF]] | ||
[[Category: Pai | |||
Latest revision as of 11:34, 25 October 2023
Crystal structure of rat xanthine oxidoreductase mutant (W335A and F336L)Crystal structure of rat xanthine oxidoreductase mutant (W335A and F336L)
Structural highlights
FunctionXDH_RAT Key enzyme in purine degradation. Catalyzes the oxidation of hypoxanthine to xanthine. Catalyzes the oxidation of xanthine to uric acid. Contributes to the generation of reactive oxygen species.[1] Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedReactive oxygen species are generated by various systems, including NADPH oxidases, xanthine oxidoreductase (XOR) and mitochondrial respiratory enzymes, and contribute to many physiological and pathological phenomena. Mammalian xanthine dehydrogenase (XDH) can be converted to xanthine oxidase (XO), which produces both superoxide anion and hydrogen peroxide in a molar ratio of about 1:3, depending upon the conditions. Here, we present a mutant of rat XOR that displays mainly XO activity with a superoxide:hydrogen peroxide production ratio of about 6:1. In the mutant, tryptophan 335, which is a component of the amino acid cluster crucial for switching from the XDH to the XO conformation, was replaced with alanine, and phenylalanine 336, which modulates FAD's redox potential through stacking interactions with the flavin cofactor, was changed to leucine. When the mutant was expressed in Sf9 cells, it was obtained in the XO form, and dithiothreitol treatment only partially restored the pyridine nucleotide-binding capacity. The crystal structure of the dithiothreitol-treated mutant at 2.3 Angstroms resolution showed the enzyme's two subunits to be quite similar, but not identical: the cluster involved in conformation-switching was completely disrupted in one subunit, but remained partly associated in the other one. The chain trace of the active site loop in this mutant is very similar to that of the bovine XO form. These results are consistent with the idea that the XDH and XO forms of the mutant are in an equilibrium that greatly favours the XO form, but the equilibrium is partly shifted towards the XDH form upon incubation with dithiothreitol. Two mutations convert mammalian xanthine oxidoreductase to highly superoxide-productive xanthine oxidase.,Asai R, Nishino T, Matsumura T, Okamoto K, Igarashi K, Pai EF, Nishino T J Biochem. 2007 Apr;141(4):525-34. Epub 2007 Feb 14. PMID:17301076[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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