title: Initiation of Escherichia coli minichromosome replication at oriC and at protein n' recognition sites. Two modes for initiating DNA synthesis in vitro.
abstract: The start sites for leading and lagging DNA strands were determined in vitro with minichromosomes as templates. Fragments from replication intermediates were analyzed by hybridization to single-stranded probes. Leading strand synthesis in the counterclockwise direction was found to originate in or close to (position 248 to -44) the minimal origin. Complementary lagging strand synthesis started several positions to the left outside of oriC. The results suggest in addition a concerted synthesis of leading and lagging strands following the dnaA directed assembly of initiation proteins at double-stranded oricC DNA (pre-replisome). In addition, DNA synthesis could initiate at protein n' recognition sequences located within and clockwise to the asnA gene. Initiation at n' sites was dependent on protein i activity, whereas leading and lagging strand initiation in the oriC region was not affected by protein i. Our results argue against an involvement of the phi X174-type primosome in the initiation of discontinuous DNA synthesis at oriC. An alternative function is suggested.
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D002876:Chromosomes, Bacterial / Q000502:physiology*; D003001:Cloning, Molecular; D004261:DNA Replication; D004262:DNA Restriction Enzymes; D004926:Escherichia coli / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D013698:Templates, Genetic
chemical_list: D001426:Bacterial Proteins; D004262:DNA Restriction Enzymes