Petit M A, Bedale W, Osipiuk J, Lu C, Rajagopalan M, McInerney P, Goodman M F, Echols H
Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720.
J Biol Chem. 1994 Sep 23;269(38):23824-9.
Replication-blocking lesions generate a signal in Escherichia coli that leads to the induction of the multigene SOS response. Among the SOS-induced genes are umuD and umuC, whose products are necessary for the increased mutation rate in induced bacteria. The mutations are likely to result from replication across the DNA lesion, and such a bypass event has been reconstituted in vitro (Rajagopalan, M., L, C., Woodgate, R., O'Donnel, M., Goodman, M. F., Echols, H. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 10777-10781). In this work, we show that the chaperone proteins promote the proper folding of UmuC protein in vitro. We treated purified and inactive UmuC with Hsp70 and Hsp60. After Hsp70 treatment, the DNA binding activity of UmuC was recovered, but the ability to promote replication across DNA lesions was not. However, lesion bypass activity was recovered upon further treatment with Hsp60. The biological significance of such a folding pathway for UmuC protein is strengthened by in vivo evidence for a role of DnaK in UV-induced mutagenesis.
复制阻断性损伤在大肠杆菌中产生一种信号，该信号导致多基因SOS应答的诱导。SOS诱导基因包括umuD和umuC，其产物是诱导细菌中突变率增加所必需的。这些突变可能是由于DNA损伤处的复制导致的，并且这种绕过事件已在体外重建（拉贾戈帕兰，M.，L，C.，伍德盖特，R.，奥唐纳，M.，古德曼，M.F.，埃科尔斯，H.（1992年）《美国国家科学院院刊》89，10777 - 10781）。在这项工作中，我们表明伴侣蛋白在体外促进UmuC蛋白的正确折叠。我们用Hsp70和Hsp60处理纯化的无活性UmuC。经Hsp70处理后，UmuC的DNA结合活性得以恢复，但促进DNA损伤处复制的能力未恢复。然而，在用Hsp60进一步处理后，损伤绕过活性得以恢复。DnaK在紫外线诱导的诱变中起作用的体内证据强化了这种UmuC蛋白折叠途径的生物学意义。
