Identification of proteins of Escherichia coli and Saccharomyces cerevisiae that specifically bind to C/C mismatches in DNA.
The pathways leading to G:C-->C:G transversions and their repair mechanisms remain uncertain. C/C and G/G mismatches arising during DNA replication are a potential source of G:C-->C:G transversions. The Escherichia coli mutHLS mismatch repair pathway efficiently corrects G/G mismatches, whereas C/C mismatches are a poor substrate. Escherichia coli must have a more specific repair pathway to correct C/C mismatches. In this study, we performed gel-shift assays to identify C/C mismatch-binding proteins in cell extracts of E. COLI: By testing heteroduplex DNA (34mers) containing C/C mismatches, two specific band shifts were generated in the gels. The band shifts were due to mismatch-specific binding of proteins present in the extracts. Cell extracts of a mutant strain defective in MutM protein did not produce a low-mobility complex. Purified MutM protein bound efficiently to the C/C mismatch-containing heteroduplex to produce the low-mobility complex. The second protein, which produced a high-mobility complex with the C/C mismatches, was purified to homogeneity, and the amino acid sequence revealed that this protein was the FabA protein of E.COLI: The high-mobility complex was not formed in cell extracts of a fabA mutant. From these results it is possible that MutM and FabA proteins are components of repair pathways for C/C mismatches in E.COLI: Furthermore, we found that Saccharomyces cerevisiae OGG1 protein, a functional homolog of E.COLI: MutM protein, could specifically bind to the C/C mismatches in DNA.
Nakahara
T
T
Department of Biological Sciences, Graduate School of Science, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Japan.
Zhang
Q M
QM
Hashiguchi
K
K
Yonei
S
S
D000251:Adenosine Triphosphatases; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D020137:Base Pair Mismatch / Q000235:genetics*; D001665:Binding Sites; D003596:Cytosine / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D043382:DNA-Formamidopyrimidine Glycosylase; D004926:Escherichia coli / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D054889:Fatty Acid Synthase, Type II; D006836:Hydro-Lyases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008970:Molecular Weight; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; D009154:Mutation / Q000235:genetics; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C074770:MutL protein, E coli; D009692:Nucleic Acid Heteroduplexes; D003596:Cytosine; D004247:DNA; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase; C468610:DNA-formamidopyrimidine glycosylase, E coli; D000251:Adenosine Triphosphatases; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; C497175:MutS protein, E coli; D006836:Hydro-Lyases; C047118:3-hydroxyacyl-(acyl-carrier-protein) dehydratase; D054889:Fatty Acid Synthase, Type II