Abstract
Taking advantage of overproduction of the uvrD protein in cells that harbor multicopy plasmids carrying the uvrD gene, we have purified protein to physical homogeneity. The purified protein possesses single-stranded DNA-dependent ATPase and ATP-dependent DNA unwinding activities, and both activities are equally inactivated by antibodies raised against DNA helicase II. Molecular weight (75,000) and chromatographic properties of the uvrD protein are also similar to those of DNA helicase II and DNA-dependent ATPase I (Richet, E., and Kohiyama, M. (1976) J. Biol. Chem. 251, 808-812; Abdel-Monem, M., Dürwald, H., and Hoffmann-Berling, H. (1977) Eur. J. Biochem. 79, 39-45). Thus, it is concluded that the uvrD protein is identical with DNA helicase II and DNA-dependent ATPase I. Expression of the uvrD gene, as assayed by DNA-dependent ATPase activity, was stimulated by exposure of bacteria to nalidixic acid or mitomycin C. No increase in ATPase activity was observed with recA mutant cells, although basic levels of DNA-dependent ATPase activity in recA+ and recA- strains were almost the same. Thus, the uvrD gene is constitutively expressed but also regulated in a recA-dependent fashion.
Highlights
Taking advantage of overproduction oftheuvrD protein in cells that harbor multicopy plasmids carrying the uvrD gene, we hpavuerified protein to physical homogeneity
Double mutants having mutations in both uurD and polA genes are inviable [13,14,15,16]. These pleiotropic phenotypes of uvrD mutants have suggested that the uurD gene product may play important roles in the processes of repair, recombination, and replication of DNA
Studies with the cloned gene revealed that the uurD gene product is a polypeptide of M, = 75,000 and possesses DNAdependent ATPase activity [18].Based on these and other findings, we have suggested that the uurD protein maybe identical with DNA-dependent ATPase 1 and possibly DNA helicase 11, which were characterized previously [19, 20]
Summary
Double mutants having mutations in both uurD and polA genes are inviable [13,14,15,16] These pleiotropic phenotypes of uvrD mutants have suggested that the uurD gene product may play important roles in the processes of repair, recombination, and replication of DNA. Studies with the cloned gene revealed that the uurD gene product is a polypeptide of M , = 75,000 and possesses DNAdependent ATPase activity [18].Based on these and other findings, we have suggested that the uurD protein maybe identical with DNA-dependent ATPase 1 and possibly DNA helicase 11, which were characterized previously [19, 20]. We present evidence that the uurD protein is identical with DNA helicase 11 and DNA-dependent ATPase I and that expression of the uurD gene is enhanced by SOS-inducing treatments
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