Replicative Deoxyribonucleic Acid Synthesis in a System Diffusible for Macromolecules

Abstract

Abstract Replicative DNA synthesis in toluene-treated Escherichia coli cells occurs in the presence of Triton X-100, a nonionic detergent. DNA replication in the detergent requires four deoxynucleoside 5'-triphosphates, Mg++, K+, and ATP, and is sensitive to sulfhydryl-blocking agents. Replicative synthesis occurs in polA1 mutants, and is temperature-sensitive in dnats mutants in Triton X-100. DNA synthesis is unaffected by the concentration of Triton X-100 present. In 1% detergent at 37° DNA polymerase I and lactate dehydrogenase are free to diffuse from the cells. Repair synthesis is inhibited by antibody to DNA polymerase I under these conditions, but ATP-dependent synthesis is not altered by such antibody. Toluene-treated cells retain their gross morphology in 1% Triton X-100. Stationary phase cells show diminished ATP-stimulated DNA synthesis compared with log stage cells after toluene treatment. In reaction mixtures containing Triton X-100, toluene-treated plateau stage cells are capable of undiminished levels of replicative synthesis. This effect does not appear to be due to increased permeability to small molecules.