Department of Biochemistry and Biomedical Sciences, McMaster University, 1200 Main Street West, Hamilton, Ontario, Canada.
DNA Repair (Amst). 2011 Jan 2;10(1):87-93. doi: 10.1016/j.dnarep.2010.10.003. Epub 2010 Nov 2.
Mismatch repair corrects errors that have escaped polymerase proofreading enhancing replication fidelity by at least two orders of magnitude. The β and PCNA sliding clamps increase the polymerase processivity during DNA replication and are important at several stages of mismatch repair. Both MutS and MutL, the two proteins that initiate the mismatch repair response, interact with β. Binding of MutS to β is important to recruit MutS and MutL to foci. Moreover, the endonuclease activity of human and yeast MutLα is stimulated by PCNA. However, the concrete functions of the processivity clamp in the repair steps preceding DNA resynthesis remain obscure. Here, we demonstrate that the C-terminal domain of MutL encompasses a bona fide β-binding motif that mediates a weak, yet specific, interaction between the two proteins. Mutation of this conserved motif correlates with defects in mismatch repair, demonstrating that the direct interaction with β is important for MutL function. The interaction between the C-terminal domain of MutL and β is conserved in both Bacillus subtilis and Escherichia coli, but the repair defects associated with mutation of this β-binding motif are more severe in the former, suggesting that this interaction may have a more prominent role in methyl-independent than methyl-directed mismatch repair systems. Together with previously published data, our work strongly suggests that β may stimulate the endonuclease activity of MutL through its direct interaction with the C-terminal domain of MutL.
错配修复纠正了逃脱聚合酶校对的错误，将复制保真度提高了至少两个数量级。β 和 PCNA 滑动夹在 DNA 复制过程中增加了聚合酶的持续合成能力，并且在错配修复的几个阶段都很重要。启动错配修复反应的两种蛋白质 MutS 和 MutL 都与β相互作用。MutS 与β的结合对于招募 MutS 和 MutL 到焦点至关重要。此外，人类和酵母 MutLα的内切核酸酶活性被 PCNA 刺激。然而，在 DNA 重新合成之前的修复步骤中，持续合成夹的具体功能仍然不清楚。在这里，我们证明 MutL 的 C 末端结构域包含一个真正的 β 结合基序，介导了两种蛋白质之间的弱但特异性相互作用。该保守基序的突变与错配修复缺陷相关，表明与β的直接相互作用对于 MutL 功能很重要。MutL 的 C 末端结构域与β之间的相互作用在枯草芽孢杆菌和大肠杆菌中都保守，但与该β结合基序突变相关的修复缺陷在前者中更为严重，这表明这种相互作用在非甲基指导的错配修复系统中可能比在甲基指导的错配修复系统中具有更突出的作用。结合之前发表的数据，我们的工作强烈表明β可能通过其与 MutL 的 C 末端结构域的直接相互作用来刺激 MutL 的内切核酸酶活性。
