Benson S A, Silhavy T J
Cell. 1983 Apr;32(4):1325-35. doi: 10.1016/0092-8674(83)90313-6.
It has been proposed that the efficient localization of the outer membrane protein LamB requires a functional signal sequence and at least two additional regions contained within the mature protein. We define these regions more precisely by deletion analysis, and we describe methods for cloning deleterious lacZ fusions onto high-copy-number plasmids and generating in-frame deletions. Analysis of the effects of a series of internal lamB deletions on the export of a LamB-LacZ hybrid protein and of the LamB protein itself indicates that necessary informational signal(s) required for localization lie at the amino-terminal end of the protein. In addition, our analysis indicates that there is a region of information close to or within the fusion joint of the largest lamB-lacZ fusion that increases the efficiency of the export process. A unique deletion that removes a protein segment from amino acid 70 to 200 appears to prevent proteolytic removal of the signal sequence. Nevertheless, the mutant protein is exported to the outer membrane.
有人提出,外膜蛋白LamB的有效定位需要一个功能性信号序列以及成熟蛋白中包含的至少两个其他区域。我们通过缺失分析更精确地定义了这些区域,并描述了将有害的lacZ融合体克隆到高拷贝数质粒上并产生框内缺失的方法。对一系列内部lamB缺失对LamB-LacZ杂合蛋白和LamB蛋白本身输出的影响分析表明,定位所需的必要信息信号位于该蛋白的氨基末端。此外,我们的分析表明,在最大的lamB-lacZ融合体的融合接头附近或内部存在一个信息区域,可提高输出过程的效率。一个独特的缺失,即从氨基酸70到200去除一个蛋白片段,似乎可防止信号序列的蛋白水解去除。然而,突变蛋白仍被输出到外膜。