The Saccharomyces cerevisiae Mcm6/2 and Mcm5/3 ATPase active sites contribute to the function of the putative Mcm2-7 'gate'.
The Mcm2-7 complex is the eukaryotic replicative helicase, a toroidal AAA(+) molecular motor that uses adenosine triphosphate (ATP) binding and hydrolysis to separate duplex DNA strands during replication. This heterohexameric helicase contains six different and essential subunits (Mcm2 through Mcm7), with the corresponding dimer interfaces forming ATPase active sites from conserved motifs of adjacent subunits. As all other known hexameric helicases are formed from six identical subunits, the function of the unique heterohexameric organization of Mcm2-7 is of particular interest. Indeed, prior work using mutations in the conserved Walker A box ATPase structural motif strongly suggests that individual ATPase active sites contribute differentially to Mcm2-7 activity. Although only a specific subset of active sites is required for helicase activity, another ATPase active site (Mcm2/5) may serve as a reversible ATP-dependent discontinuity ('gate') within the hexameric ring structure. This study analyzes the contribution that two other structural motifs, the Walker B box and arginine finger, make to each Mcm2-7 ATPase active site. Mutational analysis of these motifs not only confirms that Mcm ATPase active sites contribute unequally to activity but implicates the involvement of at least two additional active sites (Mcm5/3 and 6/2) in modulating the activity of the putative Mcm2/5 gate.
Bochman
Matthew L
ML
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
Schwacha
Anthony
A
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004270:DNA, Circular / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D064126:Minichromosome Maintenance Complex Component 3; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004270:DNA, Circular; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C069200:MCM5 protein, S cerevisiae; D009687:Nuclear Proteins; D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins; D004247:DNA; D000251:Adenosine Triphosphatases; D004265:DNA Helicases; C069199:MCM2 protein, S cerevisiae; C507761:MCM3 protein, S cerevisiae; C491207:MCM6 protein, S cerevisiae; C491088:MCM7 protein, S cerevisiae; D064126:Minichromosome Maintenance Complex Component 3; D064151:Minichromosome Maintenance Complex Component 6; D064168:Minichromosome Maintenance Complex Component 7