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Abstract:
Inactivating antibodies raised against DNA helicase I and DNA helicase II were applied to Escherichia coli DNA-replicating systems. Antibody against DNA helicase II was found to inhibit the replication of E. coli DNA, lambda phage DNA (during early and late phases), and ColE1 plasmid DNA during the elongation step. The antibody did not inhibit the replication of fd replicative form (RF) DNA, the unwinding of which is known to depend on the rep protein. Antibody against DNA helicase I failed to inhibit any of the replication processes. The replication of E. coli DNA, lambda-DNA, and ColE1 DNA is known to be initiated in a closed circle, in contrast to fd RF which is known to be initiated in a nicked circle. In conjunction with data given in the literature, our results suggest that replicative unwinding is carried out by DNA helicase II or rep protein, depending on the mechanism by which DNA replication is initiated. The concentration of DNA helicase II in E. coli, as determined by immunological methods, is 5000 to 8000 copies/cell; that of DNA helicase I is 500 to 700.
