Functional analyses of Escherichia coli MutS-beta clamp interaction in vitro and in vivo.
Escherichia coli MutS is a highly conserved mismatch repair (MMR) protein that plays a key role in recognizing DNA mismatches and the early steps of MMR. Previous studies revealed an interaction between MutS and the replicative protein beta clamp, but it remains unclear whether the interaction functions during the process of MMR. In order to provide insight into the significance of this interaction, Far Western, Surface plasmon resonance and cell survival/mutagenesis assays were used to determine its possible influences on the in vitro and in vivo properties of MutS. The results show that a quintuple mutation of MutS residues 812-816 (MutS(betaC)), or single alanine substitution mutation of MutS residues M813 or L815 completely blocks binding of MutS to beta clamp. Wild type beta clamp interferes with DNA binding by MutS. When treated with the base analog 2-aminopurine, MutS(betaC) confers more mutations and less cellular growth rate in the mutS-deficient strain than the wild type MutS. These data indicate that the MutS-beta interaction has functional consequences during MMR in E. coli.
Zhou
Ying
Y
Wuhan Institute of Virology, Chinese Academy of Sciences, Wuchang District, China.
Li
Feng
F
Chen
Yuanyuan
Y
Bi
Lijun
L
Zhang
Xian-En
XE
D015075:2-Aminopurine / Q000494:pharmacology; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D053843:DNA Mismatch Repair; D004258:DNA Polymerase III / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D051719:MutS DNA Mismatch-Binding Protein / Q000235:genetics / Q000378:metabolism*; D016296:Mutagenesis; D011485:Protein Binding / Q000235:genetics; D020349:Surface Plasmon Resonance
D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D015075:2-Aminopurine; C101139:beta subunit, DNA polymerase III; D004258:DNA Polymerase III; D051719:MutS DNA Mismatch-Binding Protein; C497175:MutS protein, E coli