Seligman L, Bailey J, Manoil C
Department of Genetics, University of Washington, Seattle 98195, USA.
J Bacteriol. 1995 May;177(9):2315-20. doi: 10.1128/jb.177.9.2315-2320.1995.
The Escherichia coli serine chemoreceptor (Tsr) is a protein with a simple topology consisting of two membrane-spanning sequences (TM1 and TM2) separating a large periplasmic domain from N-terminal and C-terminal cytoplasmic regions. We analyzed the contributions of several sequence elements to the cytoplasmic localization of the C-terminal domain by using chemoreceptor-alkaline phosphatase gene fusions. The principal findings were as follows. (i) The cytoplasmic localization of the C-terminal domain depended on TM2 but was quite tolerant of mutations partially deleting or introducing charged residues into the sequence. (ii) The basal level of C-terminal domain export was significantly higher in proteins with the wild-type periplasmic domain than in derivatives with a shortened periplasmic domain, suggesting that the large size of the wild-type domain promotes partial membrane misinsertion. (iii) The membrane insertion of deletion derivatives with a single spanning segment (TM1 or TM2) could be controlled by either an adjacent positively charged sequence or an adjacent amphipathic sequence. The results provide evidence that the generation of the Tsr membrane topology is an overdetermined process directed by an interplay of sequences promoting and opposing establishment of the normal structure.
大肠杆菌丝氨酸化学感受器（Tsr）是一种拓扑结构简单的蛋白质，由两个跨膜序列（TM1和TM2）组成，这两个序列将一个大的周质结构域与N端和C端细胞质区域分隔开来。我们通过使用化学感受器-碱性磷酸酶基因融合技术，分析了几个序列元件对C端结构域细胞质定位的贡献。主要发现如下：（i）C端结构域的细胞质定位依赖于TM2，但对部分删除或在序列中引入带电荷残基的突变具有相当的耐受性。（ii）野生型周质结构域的蛋白质中C端结构域输出的基础水平显著高于周质结构域缩短的衍生物，这表明野生型结构域的大尺寸促进了部分膜错误插入。（iii）具有单个跨膜片段（TM1或TM2）的缺失衍生物的膜插入可由相邻的带正电荷序列或相邻的两亲性序列控制。这些结果提供了证据，表明Tsr膜拓扑结构的产生是一个由促进和反对正常结构建立的序列相互作用所指导的过度决定过程。
