Blanchard A, Wroblewski H, Barroso G
Laboratoire de Microbiologie Fondamentale et Appliquée, Université de Rennes, France.
Isr J Med Sci. 1987 May;23(5):414-7.
The expression of spiralin in the transformant strain HB101 Tsp of Escherichia coli has been investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), immunodetection after electrotransfer, and crossed immunoelectrophoresis. The protein has been sought in whole cells, cytoplasm, and plasma and outer membranes of the bacterium. Untransformed E. coli cells and Spiroplasma citri cells were used as negative and positive references, respectively. Contrary to earlier claims, spiralin was detected not only in the cytoplasm of E. coli, but also in the inner and outer membranes. In addition, our results show that the protein was not secreted by the bacterium. Three forms of spiralin could be distinguished with respect to differences in apparent molecular mass: 27.5, 29.5 and 30 kilodalton (kDa). The presence of the high molecular mass polypeptide in the two membranes of E. coli invalidates the hypothesis according to which the 27.5-kDa (or 28-kDa) species is a mature form derived from the 30-kDa (or 30.5-kDa) species by the amputation of a signal sequence. Since spiralin is an acyl protein, the possibility of variation in the extent of acylation of the protein in E. coli, with subsequent variation of the apparent molecular mass, should be taken into account.
通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）、电转移后的免疫检测以及交叉免疫电泳，对螺旋体蛋白在大肠杆菌转化菌株HB101 Tsp中的表达进行了研究。在细菌的全细胞、细胞质、质膜和外膜中寻找该蛋白。未转化的大肠杆菌细胞和柑橘螺原体细胞分别用作阴性和阳性对照。与早期的说法相反，螺旋体蛋白不仅在大肠杆菌的细胞质中被检测到，在内膜和外膜中也被检测到。此外，我们的结果表明该蛋白不会被细菌分泌。根据表观分子量的差异，可以区分出三种形式的螺旋体蛋白：27.5、29.5和30千道尔顿（kDa）。大肠杆菌的两个膜中存在高分子量多肽，这使得27.5 kDa（或28 kDa）的种类是通过切除信号序列从30 kDa（或30.5 kDa）的种类衍生而来的成熟形式这一假设无效。由于螺旋体蛋白是一种酰基化蛋白，因此应考虑大肠杆菌中该蛋白酰基化程度变化以及随后表观分子量变化的可能性。
