Coordinated leading- and lagging-strand synthesis at the Escherichia coli DNA replication fork: II. Frequency of primer synthesis and efficiency of primer utilization control Okazaki fragment size Journal Article


Authors: Zechner, E. L.; Wu, C. A.; Marians, K. J.
Article Title: Coordinated leading- and lagging-strand synthesis at the Escherichia coli DNA replication fork: II. Frequency of primer synthesis and efficiency of primer utilization control Okazaki fragment size
Abstract: To investigate the role of the priming apparatus at the replication fork in determining Okazaki fragment size, the products of primer synthesis generated in vitro during rolling-circle DNA replication catalyzed by the DNA polymerase III holoenzyme, the singlestranded DNA binding protein, and the primosome on a tailed form II DNA template were isolated and characterized. The abundance of oligoribonucleotide primers and the incidence of covalent DNA chain extension of the primer population was measured under different reaction conditions known to affect the size of the products of lagging-strand DNA synthesis. These analyses demonstrated that the factors affecting Okazaki fragment length could be distinguished by either their effect on the frequency of primer synthesis or by their influence on the efficiency of initiation of DNA synthesis from primer termini. Primase and the ribonucleoside triphosphates were found to stimulate primer synthesis. The observed trend toward smaller fragment size as the concentration of these effectors was raised was apparently a direct consequence of the increased frequency of primer synthesis. The β subunit of the DNA polymerase III holoenzyme and the deoxyribonucleoside triphosphates did not alter the priming frequency; instead, the concentration of these factors influenced the ability of the lagging-strand DNA polymerase to efficiently utilize primers to initiate DNA synthesis. Maximum utilization of the available primers correlated with the lowest mean value of Okazaki fragment length. These data were used to draw general conclusions concerning the temporal order of enzymatic steps that operate during a cycle of Okazaki fragment synthesis on the lagging-strand DNA template.
Keywords: controlled study; dna binding protein; nonhuman; dna polymerase; dna replication; dna synthesis; dna; escherichia coli; catalysis; primer dna; enzyme subunit; dna, bacterial; autoradiography; electrophoresis, polyacrylamide gel; dna primase; ribonucleosides; oligoribonucleotide; dna polymerase iii; rna nucleotidyltransferases; priority journal; article; electrophoresis, agar gel; support, u.s. gov't, p.h.s.; deoxyribonucleosides
Journal Title: Journal of Biological Chemistry
Volume: 267
Issue: 6
ISSN: 0021-9258
Publisher: American Society for Biochemistry and Molecular Biology  
Date Published: 1992-02-25
Start Page: 4045
End Page: 4053
Language: English
PUBMED: 1740452
PROVIDER: scopus
DOI/URL:
Notes: Article -- Source: Scopus
Citation Impact
MSK Authors
  1. Kenneth Marians
    135 Marians
  2. Carol A. Wu
    5 Wu