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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
33
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pubmed:dateCreated |
2006-8-14
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pubmed:abstractText |
By using biochemical and structural analyses, we have investigated the catalytic mechanism of the recently discovered flavin-dependent thymidylate synthase ThyX from Paramecium bursaria chlorella virus-1 (PBCV-1). Site-directed mutagenesis experiments have identified several residues implicated in either NADPH oxidation or deprotonation activity of PBCV-1 ThyX. Chemical modification by diethyl pyrocarbonate and mass spectroscopic analyses identified a histidine residue (His53) crucial for NADPH oxidation and located in the vicinity of the redox active N-5 atom of the FAD ring system. Moreover, we observed that the conformation of active site key residues of PBCV-1 ThyX differs from earlier reported ThyX structures, suggesting structural changes during catalysis. Steady-state kinetic analyses support a reaction mechanism where ThyX catalysis proceeds via formation of distinct ternary complexes without formation of a methyl enzyme intermediate.
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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 |
Aug
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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 |
18
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pubmed:volume |
281
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
24048-57
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pubmed:dateRevised |
2006-11-15
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pubmed:meshHeading |
pubmed-meshheading:16707489-Amino Acid Sequence,
pubmed-meshheading:16707489-Amino Acid Substitution,
pubmed-meshheading:16707489-Arginine,
pubmed-meshheading:16707489-Catalysis,
pubmed-meshheading:16707489-Chlorella,
pubmed-meshheading:16707489-Conserved Sequence,
pubmed-meshheading:16707489-Crystallography, X-Ray,
pubmed-meshheading:16707489-Diethyl Pyrocarbonate,
pubmed-meshheading:16707489-Enzyme Inhibitors,
pubmed-meshheading:16707489-Flavin-Adenine Dinucleotide,
pubmed-meshheading:16707489-Glutamic Acid,
pubmed-meshheading:16707489-Histidine,
pubmed-meshheading:16707489-Kinetics,
pubmed-meshheading:16707489-Molecular Sequence Data,
pubmed-meshheading:16707489-Phycodnaviridae,
pubmed-meshheading:16707489-Sequence Alignment,
pubmed-meshheading:16707489-Structure-Activity Relationship,
pubmed-meshheading:16707489-Substrate Specificity,
pubmed-meshheading:16707489-Thymidylate Synthase
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pubmed:year |
2006
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pubmed:articleTitle |
Catalytic mechanism and structure of viral flavin-dependent thymidylate synthase ThyX.
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pubmed:affiliation |
CNRS, UMR 7645, Laboratory of Optics and Biosciences, Ecole Polytechnique, 91128 Palaiseau, France.
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pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
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