Makovets S, Doronina V A, Murray N E
Institute of Cell and Molecular Biology, Darwin Building, King's Buildings, University of Edinburgh, Edinburgh EH9 3JR, United Kingdom.
Proc Natl Acad Sci U S A. 1999 Aug 17;96(17):9757-62. doi: 10.1073/pnas.96.17.9757.
ClpXP-dependent proteolysis has been implicated in the delayed detection of restriction activity after the acquisition of the genes (hsdR, hsdM, and hsdS) that specify EcoKI and EcoAI, representatives of two families of type I restriction and modification (R-M) systems. Modification, once established, has been assumed to provide adequate protection against a resident restriction system. However, unmodified targets may be generated in the DNA of an hsd(+) bacterium as the result of replication errors or recombination-dependent repair. We show that ClpXP-dependent regulation of the endonuclease activity enables bacteria that acquire unmodified chromosomal target sequences to survive. In such bacteria, HsdR, the polypeptide of the R-M complex essential for restriction but not modification, is degraded in the presence of ClpXP. A mutation that blocks only the modification activity of EcoKI, leaving the cell with approximately 600 unmodified targets, is not lethal provided that ClpXP is present. Our data support a model in which the HsdR component of a type I restriction endonuclease becomes a substrate for proteolysis after the endonuclease has bound to unmodified target sequences, but before completion of the pathway that would result in DNA breakage.
ClpXP 依赖性蛋白酶解作用与在获得指定 EcoKI 和 EcoAI（I 型限制与修饰（R-M）系统两个家族的代表）的基因（hsdR、hsdM 和 hsdS）后限制活性的延迟检测有关。一旦建立修饰作用，人们认为它能为抵御常驻限制系统提供充分保护。然而，由于复制错误或重组依赖性修复，hsd(+)细菌的 DNA 中可能会产生未修饰的靶标。我们表明，ClpXP 对核酸内切酶活性的调控使获得未修饰染色体靶标序列的细菌得以存活。在这类细菌中，R-M 复合物中对限制而非修饰至关重要的多肽 HsdR，在 ClpXP 存在的情况下会被降解。只要存在 ClpXP，仅阻断 EcoKI 修饰活性、使细胞带有约 600 个未修饰靶标的突变就不会致死。我们的数据支持这样一种模型，即 I 型限制核酸内切酶的 HsdR 组分在核酸内切酶与未修饰靶标序列结合后，但在导致 DNA 断裂的途径完成之前，会成为蛋白酶解的底物。
