Biochemical activities of the BOB1 mutant in Methanobacterium thermoautotrophicum MCM.
Minichromosomal maintenance proteins (MCMs) are considered to be the replicative helicase. Methanobacterium thermoautotrophicum has a single MCM gene (mtMCM). The crystal structure of the mtMCM N-terminal region is a double hexamer. Structure-guided sequence alignment indicates a structural conservation of this fragment across archaeal and eukaryotic MCMs. The mtMCM structure was successfully used to analyze a Saccharomyces cerevisiae MCM5 mutant, called BOB1, which contains a single residue change from Pro to Leu and bypasses a kinase normally required for initiation of DNA replication. A domain-push model was proposed to explain the BOB1 bypass activity. Here we investigate the effects of BOB1 mutation on the biochemical activities of mtMCM. Surprisingly, the BOB1 mutation (P62L) had a major effect on the helicase activity but had no significant impact on DNA binding and ATPase activities. These results will contribute to a more detailed understanding of the BOB1 bypass activity and other aspects of DNA replication control.
Fletcher
Ryan J
RJ
Molecular and Computational Biology, University of Southern California, Los Angeles, California 90089, USA.
Chen
Xiaojiang S
XS
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D019943:Amino Acid Substitution / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D018797:Cell Cycle Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular; D017124:Conserved Sequence / Q000235:genetics; D004265:DNA Helicases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D019641:DNA, Archaeal / Q000235:genetics / Q000378:metabolism; D004272:DNA, Mitochondrial / Q000235:genetics / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D019143:Evolution, Molecular; D007930:Leucine / Q000235:genetics; D017014:Methanobacterium / Q000737:chemistry* / Q000235:genetics*; D026341:Minichromosome Maintenance 1 Protein / Q000378:metabolism*; D011392:Proline / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification
D018797:Cell Cycle Proteins; D019641:DNA, Archaeal; D004272:DNA, Mitochondrial; D004277:DNA, Single-Stranded; C069200:MCM5 protein, S cerevisiae; D026341:Minichromosome Maintenance 1 Protein; D029701:Saccharomyces cerevisiae Proteins; D011392:Proline; D000251:Adenosine Triphosphatases; D004265:DNA Helicases; D007930:Leucine