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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Different modes of SecY-SecA interactions revealed by site-directed in vivo photo-cross-linking
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Different modes of SecY-SecA interactions revealed by site-directed in vivo photo-cross-linking

机译:定点体内光交联揭示了SecY-SecA相互作用的不同模式

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While the SecA ATPase drives protein translocation across the bacteria[ cytoplasmic membrane by interacting with the SecYEG translocon, molecular details of SecA-SecY interaction remain poorly understood. Here, we studied SecY-SecA interaction by using an in vivo site-directed cross-linking technique developed by Schultz and coworkers [Chin, J. W., Martin, A. B., King, D. S., Wang, L., Schultz, P. G. (2002) Proc. Natl. Acad. Sci. USA 99:11020-11024 and Chin, J. W., Schultz, P. G. (2002) ChemBioChem 3:1135-1137]. Benzoyl-phenylalanine introduced into specific SecY positions at the second, fourth, fifth, and sixth cytoplasmic domains allowed UV cross-linking with Ser-A. Cross-linked products exhibited two distinct electrophoretic mobilities. SecA cross-linking at the most C-terminal cytoplasmic region (C6) was specifically enhanced in the presence of NaN3, which arrests the ATPase cycle, and this enhancement was canceled by cis placement of some secY mutations that affect SecY-SecA cooperation. In vitro experiments showed directly that SecA approaches C6 when it is engaging in ATP-dependent preprotein translocation. On the basis of these findings, we propose that the C6 tail of SecY interacts with the working form of SecA, whereas C4-C5 loops may offer constitutive SecA-binding sites.
机译:虽然SecA ATPase通过与SecYEG转运子相互作用而驱动蛋白在细菌细胞质膜中移位,但对SecA-SecY相互作用的分子细节仍知之甚少。在这里,我们使用舒尔茨及其同事[Chin,JW,Martin,AB,King,DS,Wang,L.,Schultz,PG(2002)Proc。 。 Natl。学院科学USA 99:11020-11024和Chin,J.W.,Schultz,PG。(2002)ChemBioChem 3:1135-1137]。在第二,第四,第五和第六个胞质结构域中的特定SecY位置引入苯甲酰基-苯丙氨酸可使UV与Ser-A交联。交联产物表现出两种不同的电泳迁移率。在存在NaN3的情况下,SecA在最C端细胞质区域(C6)的交联被特别增强,这会阻止ATPase循环,并且这种顺式作用会影响SecY-SecA合作的某些secY突变被抵消。体外实验直接表明,SecA在参与ATP依赖的前蛋白易位时接近C6。根据这些发现,我们建议SecY的C6尾部与SecA的工作形式相互作用,而C4-C5环可能提供组成型SecA结合位点。

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