phiX174 DNA-dependent DNA synthesis in vitro: requirement for P1 ban protein in dnaB mutant extracts of Escherichia coli
- PMID: 670
- PMCID: PMC433105
- DOI: 10.1073/pnas.72.10.3907
phiX174 DNA-dependent DNA synthesis in vitro: requirement for P1 ban protein in dnaB mutant extracts of Escherichia coli
Abstract
Ammonium sulfate fractionation of crude extracts of E. coli yields a soluble enzyme fraction (about 25-fold purification) that catalyzes the conversion of phiX174 single-stranded DNA to duplex DNA. The reaction is rifampicin-resistant, requires single-stranded DNA, Mg++, deoxynucleoside triphosphates, and ATP, and is stimulated by KCl. Such soluble enzyme fractions were prepared from E. coli strains carrying the prophage mutant P1bac, in which the viral dnaB analog (ban) protein is expressed constitutively, or P1bacban, in which the expression of ban protein is prevented. DNA-synthesizing activity of ban protein containing fractions from wild-type or dnaB(P1bac) lysogens was more temperature-resistant than that from E. coli containing only wild-type dnaB protein, whereas that from dnaB(P1bacban) lysogens of dnaB cells was extremely thermolabile. It is suggested that the temperature-resistant DNA synthesis with fractions from P1bac lysogens is mediated by the P1 ban protein.
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