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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
14
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pubmed:dateCreated |
2000-5-8
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pubmed:abstractText |
The mechanism of the 5'-2-deoxyribose-5-phosphate lyase reaction catalyzed by mammalian DNA beta-polymerase (beta-pol) was investigated using a cross-linking methodology in combination with mass spectrometric analyses. The approach included proteolysis of the covalently cross-linked protein-DNA complex with trypsin, followed by isolation, peptide mapping, and mass spectrometric and tandem mass spectrometric analyses. The 8-kDa domain of beta-pol was covalently cross-linked to a 5'-2-deoxyribose-5-phosphate-containing DNA substrate by sodium borohydride reduction. Using tandem mass spectrometry, the location of the DNA adduct on the 8-kDa domain was unequivocally determined to be at the Lys(72) residue. No additional amino acid residues were found as minor cross-linked species. These data allow assignment of Lys(72) as the sole Schiff base nucleophile in the 8-kDa domain of beta-pol. These results provide the first direct evidence in support of a catalytic mechanism involving nucleophilic attack by Lys(72) at the abasic site.
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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 |
Apr
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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 |
7
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pubmed:volume |
275
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
10463-71
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pubmed:dateRevised |
2006-11-15
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pubmed:meshHeading |
pubmed-meshheading:10744736-Amino Acid Sequence,
pubmed-meshheading:10744736-Binding Sites,
pubmed-meshheading:10744736-Borohydrides,
pubmed-meshheading:10744736-Cross-Linking Reagents,
pubmed-meshheading:10744736-DNA,
pubmed-meshheading:10744736-DNA Polymerase beta,
pubmed-meshheading:10744736-Lysine,
pubmed-meshheading:10744736-Mass Spectrometry,
pubmed-meshheading:10744736-Models, Molecular,
pubmed-meshheading:10744736-Molecular Sequence Data,
pubmed-meshheading:10744736-Molecular Weight,
pubmed-meshheading:10744736-Nucleic Acid Conformation,
pubmed-meshheading:10744736-Oligodeoxyribonucleotides,
pubmed-meshheading:10744736-Peptide Mapping,
pubmed-meshheading:10744736-Protein Conformation,
pubmed-meshheading:10744736-Schiff Bases,
pubmed-meshheading:10744736-Trypsin
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pubmed:year |
2000
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pubmed:articleTitle |
Mapping of the 5'-2-deoxyribose-5-phosphate lyase active site in DNA polymerase beta by mass spectrometry.
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pubmed:affiliation |
Laboratory of Structural Biology, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
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pubmed:publicationType |
Journal Article
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