The C-terminal domain of the Pseudomonas secretin XcpQ forms oligomeric rings with pore activity

被引:70
作者
Brok, R
Van Gelder, P
Winterhalter, M
Ziese, U
Koster, AJ
de Cock, H
Koster, M
Tommassen, J
Bitter, W
机构
[1] Univ Utrecht, Dept Mol Microbiol, NL-3584 CH Utrecht, Netherlands
[2] Univ Utrecht, Dept Mol Cell Biol, NL-3584 CH Utrecht, Netherlands
[3] Univ Basel, Biozentrum, Dept Biophys, Basel, Switzerland
关键词
secretin; type II protein secretion; general secretory pathway; outer membrane; Pseudomonas;
D O I
10.1006/jmbi.1999.3340
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The Pseudomonas secretin XcpQ forms an oligomeric complex, which is involved in the translocation of proteins across the outer membrane via the type II secretion pathway. Pseudomonas aeruginosa produces only small amounts of this complex, 50 to 100 copies per bacterium, and overexpression is lethal to these cells. However, overexpression of Pseudomonas alcaligenes XcpQ could be achieved in the P. alcaligenes mutant strain 537. Protease protection experiments with P. alcaligenes XcpQ showed that the C-terminal domain of XcpQ, which is conserved in all the different members of the secretin family, is largely resistant to proteinase K. This protease-resistant fragment is embedded in the membrane and remains a stable complex, indicating that this domain is involved in complex formation. Both the intact and the protease-protected XcpQ complex showed a tendency to form two-dimensional crystal-like structures. Electron microscopic analysis of these structures showed that the overall oligomeric rings of the intact and of the protease-resistant complex are highly similar. The central cavity of the intact XcpQ complex contains structured mass. Both the intact and the protease-protected XcpQ complex showed pore-forming activity in planar lipid bilayers, consistent with their role as a translocation channel. However, the single-channel conductances observed were not uniform. Together, these results demonstrate that the C-terminal secretin homology domain of XcpQ is the structural domain that forms the channel through which macromolecules are being transported. (C) 1999 Academic Press.
引用
收藏
页码:1169 / 1179
页数:11
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