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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
39
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pubmed:dateCreated |
1995-11-6
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pubmed:abstractText |
The roles of active site residues His54, Phe94, Lys183, and His220 in the Streptomyces rubiginosus D-xylose isomerase were probed by site-directed mutagenesis. The kinetic properties and crystal structures of the mutant enzymes were characterized. The pH dependence of diethylpyrocarbonate modification of His54 suggests that His54 does not catalyze ring-opening as a general acid. His54 appears to be involved in anomeric selection and stabilization of the acyclic transition state by hydrogen bonding. Phe94 stabilizes the acyclic-extended transition state directly by hydrophobic interactions and/or indirectly by interactions with Trp137 and Phe26. Lys183 and His220 mutants have little or no activity and the structures of these mutants with D-xylose reveal cyclic alpha-D-xylopyranose. Lys183 functions structurally by maintaining the position of Pro187 and Glu186 and catalytically by interacting with acyclic-extended sugars. His220 provides structure for the M2-metal binding site with properties which are necessary for extension and isomerization of the substrate. A second M2 metal binding site (M2') is observed at a relatively lower occupancy when substrate is added consistent with the hypothesis that the metal moves as the hydride is shifted on the extended substrate.
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pubmed:grant |
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pubmed:language |
eng
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pubmed:journal |
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pubmed:citationSubset |
IM
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pubmed:chemical |
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pubmed:status |
MEDLINE
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pubmed:month |
Sep
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pubmed:issn |
0021-9258
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pubmed:author |
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pubmed:issnType |
Print
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pubmed:day |
29
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pubmed:volume |
270
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
22895-906
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pubmed:dateRevised |
2011-11-17
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pubmed:meshHeading |
pubmed-meshheading:7559425-Aldose-Ketose Isomerases,
pubmed-meshheading:7559425-Amino Acid Sequence,
pubmed-meshheading:7559425-Base Sequence,
pubmed-meshheading:7559425-Binding Sites,
pubmed-meshheading:7559425-Carbohydrate Epimerases,
pubmed-meshheading:7559425-Cloning, Molecular,
pubmed-meshheading:7559425-Crystallography, X-Ray,
pubmed-meshheading:7559425-DNA Primers,
pubmed-meshheading:7559425-Escherichia coli,
pubmed-meshheading:7559425-Histidine,
pubmed-meshheading:7559425-Kinetics,
pubmed-meshheading:7559425-Lysine,
pubmed-meshheading:7559425-Models, Molecular,
pubmed-meshheading:7559425-Molecular Sequence Data,
pubmed-meshheading:7559425-Mutagenesis, Site-Directed,
pubmed-meshheading:7559425-Phenylalanine,
pubmed-meshheading:7559425-Point Mutation,
pubmed-meshheading:7559425-Polymerase Chain Reaction,
pubmed-meshheading:7559425-Protein Conformation,
pubmed-meshheading:7559425-Recombinant Proteins,
pubmed-meshheading:7559425-Streptomyces,
pubmed-meshheading:7559425-Tryptophan
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pubmed:year |
1995
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pubmed:articleTitle |
Probing the roles of active site residues in D-xylose isomerase.
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pubmed:affiliation |
Department of Food Science, Cornell University, Ithaca, New York 14853, USA.
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pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, U.S. Gov't, P.H.S.,
Research Support, Non-U.S. Gov't
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