Coupling DNA unwinding activity with primer synthesis in the bacteriophage T4 primosome.
The unwinding and priming activities of the bacteriophage T4 primosome, which consists of a hexameric helicase (gp41) translocating 5' to 3' and an oligomeric primase (gp61) synthesizing primers 5' to 3', have been investigated on DNA hairpins manipulated by a magnetic trap. We find that the T4 primosome continuously unwinds the DNA duplex while allowing for primer synthesis through a primosome disassembly mechanism or a new DNA looping mechanism. A fused gp61-gp41 primosome unwinds and primes DNA exclusively via the DNA looping mechanism. Other proteins within the replisome control the partitioning of these two mechanisms by disfavoring primosome disassembly, thereby increasing primase processivity. In contrast to T4, priming in bacteriophage T7 and Escherichia coli involves discrete pausing of the primosome and dissociation of the primase from the helicase, respectively. Thus nature appears to use several strategies to couple the disparate helicase and primase activities within primosomes.
Manosas
Maria
M
Université Pierre et Marie Curie Paris, Université Paris Diderot, Centre National de la Recherche Scientifique, France.
Spiering
Michelle M
MM
Zhuang
Zhihao
Z
Benkovic
Stephen J
SJ
Croquette
Vincent
V
D017122:Bacteriophage T4 / Q000201:enzymology* / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D019915:DNA Primase / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers / Q000096:biosynthesis*; D004261:DNA Replication; D008954:Models, Biological; D012313:RNA / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D017931:DNA Primers; C034477:RNA primers; D011993:Recombinant Fusion Proteins; D012313:RNA; D004247:DNA; D019915:DNA Primase; D004265:DNA Helicases