Sakaguchi M, Tomiyoshi R, Kuroiwa T, Mihara K, Omura T
Department of Molecular Biology, Graduate School of Medical Science, Fukuoka, Japan.
Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):16-9. doi: 10.1073/pnas.89.1.16.
The signal sequence of secretory proteins and the signal-anchor sequence of type II membrane proteins initiate the translocation of the following polypeptide segments, whereas the signal-anchor sequence of cytochrome P-450-type membrane proteins mediates the membrane insertion of the polypeptide via a signal-recognition particle-dependent mechanism but does not lead to the translocation of the following C-terminal sequences. To establish the structural requirements for the function of signal and signal-anchor sequences, we constructed chimeric proteins containing artificial topogenic sequences in which the N-terminal net charge and the length of the hydrophobic segment were systematically altered. Utilizing an in vitro translation-translocation system, we found that hydrophobic segments consisting of 7-10 leucine residues functioned as signal sequences whereas segments with 12-15 leucine residues showed different topogenic functions, behaving as signal sequences or P-450-type signal-anchor sequences, depending on the N-terminal charge. From these observations, we propose that the function of N-terminal topogenic sequences depends on a balance between the N-terminal charge and the length of the following hydrophobic segment.
分泌蛋白的信号序列和II型膜蛋白的信号锚定序列启动后续多肽片段的转运，而细胞色素P-450型膜蛋白的信号锚定序列通过依赖信号识别颗粒的机制介导多肽的膜插入，但不会导致后续C末端序列的转运。为了确定信号序列和信号锚定序列功能的结构要求，我们构建了包含人工拓扑序列的嵌合蛋白，其中N端净电荷和疏水片段的长度被系统地改变。利用体外翻译-转运系统，我们发现由7-10个亮氨酸残基组成的疏水片段作为信号序列发挥作用，而具有12-15个亮氨酸残基的片段表现出不同的拓扑功能，根据N端电荷的不同，表现为信号序列或P-450型信号锚定序列。基于这些观察结果，我们提出N端拓扑序列的功能取决于N端电荷与后续疏水片段长度之间的平衡。
