Source:http://linkedlifedata.com/resource/pubmed/id/11401542
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Predicate | Object |
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
24
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pubmed:dateCreated |
2001-6-12
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pubmed:abstractText |
Dihydroorotase plays a key role in pyrimidine biosynthesis by catalyzing the reversible interconversion of carbamoyl aspartate to dihydroorotate. Here we describe the three-dimensional structure of dihydroorotase from Escherichia coli determined and refined to 1.7 A resolution. Each subunit of the homodimeric enzyme folds into a "TIM" barrel motif with eight strands of parallel beta-sheet flanked on the outer surface by alpha-helices. Unexpectedly, each subunit contains a binuclear zinc center with the metal ions separated by approximately 3.6 A. Lys 102, which is carboxylated, serves as a bridging ligand between the two cations. The more buried or alpha-metal ion in subunit I is surrounded by His 16, His 18, Lys 102, Asp 250, and a solvent molecule (most likely a hydroxide ion) in a trigonal bipyramidal arrangement. The beta-metal ion, which is closer to the solvent, is tetrahedrally ligated by Lys 102, His 139, His 177, and the bridging hydroxide. L-Dihydroorotate is observed bound to subunit I, with its carbonyl oxygen, O4, lying 2.9 A from the beta-metal ion. Important interactions for positioning dihydroorotate into the active site include a salt bridge with the guanidinium group of Arg 20 and various additional electrostatic interactions with both protein backbone and side chain atoms. Strikingly, in subunit II, carbamoyl L-aspartate is observed binding near the binuclear metal center with its carboxylate side chain ligating the two metals and thus displacing the bridging hydroxide ion. From the three-dimensional structures of the enzyme-bound substrate and product, it has been possible to propose a unique catalytic mechanism for dihydroorotase. In the direction of dihydroorotate hydrolysis, the bridging hydroxide attacks the re-face of dihydroorotate with general base assistance by Asp 250. The carbonyl group is polarized for nucleophilic attack by the bridging hydroxide through a direct interaction with the beta-metal ion. During the cyclization of carbamoyl aspartate, Asp 250 initiates the reaction by abstracting a proton from N3 of the substrate. The side chain carboxylate of carbamoyl aspartate is polarized through a direct electrostatic interaction with the binuclear metal center. The ensuing tetrahedral intermediate collapses with C-O bond cleavage and expulsion of the hydroxide which then bridges the binuclear metal center.
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pubmed:grant | |
pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Aryldialkylphosphatase,
http://linkedlifedata.com/resource/pubmed/chemical/Aspartic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Carbamyl Phosphate,
http://linkedlifedata.com/resource/pubmed/chemical/Dihydroorotase,
http://linkedlifedata.com/resource/pubmed/chemical/Esterases,
http://linkedlifedata.com/resource/pubmed/chemical/Lysine,
http://linkedlifedata.com/resource/pubmed/chemical/Orotic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Zinc
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pubmed:status |
MEDLINE
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pubmed:month |
Jun
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pubmed:issn |
0006-2960
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
19
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pubmed:volume |
40
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
6989-97
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pubmed:dateRevised |
2007-11-14
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pubmed:meshHeading |
pubmed-meshheading:11401542-Amino Acid Sequence,
pubmed-meshheading:11401542-Aryldialkylphosphatase,
pubmed-meshheading:11401542-Aspartic Acid,
pubmed-meshheading:11401542-Binding Sites,
pubmed-meshheading:11401542-Carbamyl Phosphate,
pubmed-meshheading:11401542-Catalysis,
pubmed-meshheading:11401542-Crystallography, X-Ray,
pubmed-meshheading:11401542-Dihydroorotase,
pubmed-meshheading:11401542-Dimerization,
pubmed-meshheading:11401542-Escherichia coli,
pubmed-meshheading:11401542-Esterases,
pubmed-meshheading:11401542-Humans,
pubmed-meshheading:11401542-Lysine,
pubmed-meshheading:11401542-Molecular Sequence Data,
pubmed-meshheading:11401542-Orotic Acid,
pubmed-meshheading:11401542-Structure-Activity Relationship,
pubmed-meshheading:11401542-Zinc
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pubmed:year |
2001
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pubmed:articleTitle |
Molecular structure of dihydroorotase: a paradigm for catalysis through the use of a binuclear metal center.
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pubmed:affiliation |
Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706, USA.
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pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, U.S. Gov't, P.H.S.,
Research Support, U.S. Gov't, Non-P.H.S.
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