Summary
An Escherichia coli K12 dnaB dnaC mutant was constructed by P1 transduction of the dnaC allele into a dnaB recipient strain dnaB dnaC transductants were discriminated from dnaB mutants by their inability to grow at 40° C after lysogenization with phage P1bac. The dnaB dnaC mutant character was verified by 1. P1 transduction, and 2. by in vitro complementation with dnaB and dnaC wild type protein fractions.
DNA synthesis was studied in strains containing dnaB, dnaC, or dnaB dnaC alleles in an otherwise uniform genetic background with the dnaB character either unsuppressed or suppressed by P1bac prophage. Degradation at 42° C of [3H]-thymidine pulselabeled DNA in dnaB and dnaB dnaC mutants is suppressed by P1bac. However, unlike the dnaC mutant, the P1bac lysogen of the dnaB dnaC mutant exhibits an abrupt cessation of DNA synthesis and less residual cell divisions at 42° C indicating an inhibition of DNA chain elongation rather than a defect in DNA initiation. It is suggested that denaturation of the dnaB protein affects the dnaC function.
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Ahmad, S.I., Pritchard, R.H.: A map of four genes specifying enzymes involved in catabolism of nucleosides and deoxynucleosides in Escherichia coli. Molec. gen. Genet. 104, 351–359 (1969)
Beyersmann, D., Messer, W., Schlicht, M.: Mutants of Escherichia coli B/r defective in deoxyribonucleic acid initiation: dnaI, a new gene for replication. J. Bact. 118, 783–789 (1974)
Beyersmann, D., Schlicht, M., Schuster, H.: Temperature-sensitive initiation of DNA replication in a mutant of Escherichia coli K12. Molec. gen. Genet. 111, 145–158 (1971)
Bouché, J.-P., Zechel, K., Kornberg, A.: dnaG gene product, a rifampicin-resistant RNA polymerase, initiates the conversion of a single-stranded coliphage DNA to its duplex replicative form. J. biol. Chem. 250, 5995–6001 (1975)
D'Ari, R., Jaffé-Brachet, A., Touati-Schwartz, D., Yarmolinsky, M.B.: A dnaB analog specified by bacteriophage P1. J. molec. Biol. 94, 341–366 (1975)
Geider, K.: Molecular aspects of DNA replication in Escherichia coli systems. Curr. Top. Microbiol. Immunol. 74, 55–112 (1976)
Mikolajczyk, M., Schuster, H.: The fate of newly made DNA in Escherichia coli mutants thermosensitive in DNA synthesis. Molec. gen. Genet. 110, 272–288 (1971)
Ogawa, T.: Analysis of the dnaB function of Escherichia coli K12 and the dnaB-like function of P1 prophage. J. molec. Biol. 94, 327–340 (1975)
Scheckman, R., Weiner, A., Kornberg, A.: Multienzyme systems of DNA replication. Proteins required for chromosome replication are resolved with the aid of a simple viral DNA template. Science 186, 987–993 (1974)
Schekman, R., Weiner, J.H., Weiner, A., Kornberg, A.: Ten proteins required for conversion of ϕX174 single-stranded DNA to duplex form in vitro. J. biol. Chem. 250, 5859–5865 (1975)
Schuster, H., Mikolajczyk, M., Rohrschneider, J., Geschke, B.: ϕX174 DNA-dependent DNA synthesis in vitro: Requirement for P1 ban protein in dnaB mutant extracts of Escherichia coli. Proc. nat. Acad. Sci. (Wash.) 72, 3907–3911 (1975)
Vogel, H.J., Bonner, D.M.: Acetylornithinase of Escherichia coli: Partial purification and some properties. J. biol. Chem. 218, 97–106 (1956)
Wechsler, J.A.: Genetic and phenotypic characterization of dnaC mutations. J. Bact. 121, 594–599 (1975)
Wechsler, J.A., Gross, J.D.: Escherichia coli mutants temperature-sensitive for DNA synthesis. Molec. gen. Genet. 113, 273–284 (1971)
Wechsler, J.A., Nüsslein, V., Otto, B., Klein, A., Bonhoeffer, F., Herrmann, R., Gloger, L., Schaller, H.: Isolation and characterization of thermosensitive Escherichia coli mutants defective in deoxyribonucleic acid replication. J. Bact. 113, 1381–1388 (1973)
Wickner, S., Hurwitz, J.: Interaction of Escherichia coli dnaB and dnaC(D) gene products in vitro. Proc. nat. Acad. Sci. (Wash.) 72, 921–925 (1975)
Wickner, S., Hurwitz, J.: Involvement of Escherichia coli dnaZ gene product in DNA elongation in vitro. Proc. nat. Acad. Sci. (Wash.) 73, 1053–1057 (1976)
Wickner, S., Wright, M., Hurwitz, J.: Studies on in vitro DNA synthesis. Purification of the dnaG gene product from Escherichia coli. Proc. nat. Acad. Sci. (Wash.) 70, 1613–1618 (1973)
Young, II, E.T., Sinsheimer, R.L.: Vegetative bacteriophage λ DNA. I. Infectivity in a spheroplast assay. J. molec. Biol. 30, 147–164 (1967)
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Schuster, H., Schlicht, M., Lanka, E. et al. DNA synthesis in an Escherichia coli dnaB dnaC mutant. Molec. Gen. Genet. 151, 11–16 (1977). https://doi.org/10.1007/BF00446907
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DOI: https://doi.org/10.1007/BF00446907