Evans Lewis D B, Stafford Graham P, Ahmed Sangita, Fraser Gillian M, Hughes Colin
Department of Pathology, Cambridge University, Tennis Court Road, Cambridge CB2 1QP, United Kingdom.
Proc Natl Acad Sci U S A. 2006 Nov 14;103(46):17474-9. doi: 10.1073/pnas.0605197103. Epub 2006 Nov 6.
Assembly of the bacterial flagellar filament requires a type III export pathway for ordered delivery of structural subunits from the cytosol to the cell surface. This is facilitated by transient interaction with chaperones that protect subunits and pilot them to dock at the membrane export ATPase complex. We reveal that the essential export protein FliJ has a novel chaperone escort function in the pathway, specifically recruiting unladen chaperones for the minor filament-class subunits of the filament cap and hook-filament junction substructures. FliJ did not recognize unchaperoned subunits or chaperone-subunit complexes, and it associated with the membrane ATPase complex, suggesting a function postdocking. Empty chaperones that were recruited by FliJ in vitro were efficiently captured from FliJ-chaperone complexes by cognate subunits. FliJ and subunit bound to the same region on the target chaperone, but the cognate subunit had a approximately 700-fold greater affinity for chaperone than did FliJ. The data show that FliJ recruits chaperones and transfers them to subunits, and indicate that this is driven by competition for a common binding site. This escort mechanism provides a means by which free export chaperones can be cycled after subunit release, establishing a new facet of the secretion process. As FliJ does not escort the chaperone for the major filament subunit, cycling may offer a mechanism for export selectivity and thus promote assembly of the junction and cap substructures required for initiation of flagellin polymerization.
细菌鞭毛丝的组装需要III型输出途径，以便将结构亚基从胞质溶胶有序地输送到细胞表面。这通过与伴侣蛋白的短暂相互作用得以促进，伴侣蛋白保护亚基并引导它们停靠在膜输出ATP酶复合物上。我们发现，必需的输出蛋白FliJ在该途径中具有一种新的伴侣蛋白护送功能，具体而言，它为鞭毛帽和钩-丝连接亚结构的次要丝类亚基招募空载的伴侣蛋白。FliJ不识别未结合伴侣蛋白的亚基或伴侣蛋白-亚基复合物，并且它与膜ATP酶复合物相关联，这表明其功能在停靠之后。在体外被FliJ招募的空载伴侣蛋白被同源亚基从FliJ-伴侣蛋白复合物中有效地捕获。FliJ和亚基结合在目标伴侣蛋白的同一区域，但同源亚基对伴侣蛋白的亲和力比对FliJ的亲和力大约高700倍。数据表明，FliJ招募伴侣蛋白并将它们转移给亚基，并且表明这是由对共同结合位点的竞争驱动的。这种护送机制提供了一种方式，通过该方式，游离的输出伴侣蛋白可以在亚基释放后循环利用，从而确立了分泌过程的一个新方面。由于FliJ不护送主要丝亚基的伴侣蛋白循环利用可能提供了一种输出选择性的机制，从而促进鞭毛蛋白聚合起始所需的连接和帽亚结构的组装。
