Hemilä H, Palva A, Paulin L, Arvidson S, Palva I
Genesit Ltd., Helsinki, Finland.
J Bacteriol. 1990 Sep;172(9):5052-63. doi: 10.1128/jb.172.9.5052-5063.1990.
We have cloned the operon coding for the Bacillus subtilis S complex, which has been proposed to be a component in protein secretion machinery. A lambda gt10 library of B. subtilis was screened with antiserum directed against the Staphylococcus aureus membrane-bound ribosome protein complex, which is homologous to the B. subtilis S complex. Two positive overlapping lambda clones were sequenced. The S-complex operon, 5 kilobases in size, was shown to contain four open reading frames and three putative promoters, which are located upstream of the first, the third, and the last gene. The four proteins encoded by the operon are 42, 36, 48, and 50 kilodaltons in size. All of these proteins were recognized by antisera separately raised against each protein of the S. aureus membrane-bound ribosome protein and B. subtilis S complexes, thus verifying the S-complex identity of the lambda clones. Sequence analysis revealed that all four proteins of the B. subtilis S complex are homologous to the four subunits of the human pyruvate dehydrogenase (PDH). Also, the N terminus of the 48-kilodalton protein was found to have 70% amino acid identity with the N-terminal 211 amino acids, determined so far, from the E2 subunit of B. stearothermophilus PDH. Furthermore, chromosomal mapping of the S-complex operon gave a linkage to a marker gene located close to the previously mapped B. subtilis PDH genes. Thus, the S complex is evidently identical to the B. subtilis PDH, which has been shown to contain four subunits with molecular weights very similar to those of the S complex. Therefore, we propose that the S complex is not a primary component of protein secretion.
我们已经克隆了编码枯草芽孢杆菌S复合物的操纵子，该复合物被认为是蛋白质分泌机制的一个组成部分。用针对金黄色葡萄球菌膜结合核糖体蛋白复合物的抗血清筛选枯草芽孢杆菌的λgt10文库，该复合物与枯草芽孢杆菌S复合物同源。对两个阳性重叠λ克隆进行了测序。大小为5千碱基的S复合物操纵子显示包含四个开放阅读框和三个推定的启动子，它们位于第一个、第三个和最后一个基因的上游。该操纵子编码的四种蛋白质大小分别为42、36、48和50千道尔顿。所有这些蛋白质都被分别针对金黄色葡萄球菌膜结合核糖体蛋白和枯草芽孢杆菌S复合物的每种蛋白质产生的抗血清所识别，从而验证了λ克隆的S复合物身份。序列分析表明，枯草芽孢杆菌S复合物的所有四种蛋白质与人丙酮酸脱氢酶（PDH）的四个亚基同源。此外，发现48千道尔顿蛋白质的N末端与嗜热脂肪芽孢杆菌PDH的E2亚基迄今确定 的N末端211个氨基酸具有70%的氨基酸同一性。此外，S复合物操纵子的染色体定位与一个标记基因连锁，该标记基因位于先前定位的枯草芽孢杆菌PDH基因附近。因此，S复合物显然与枯草芽孢杆菌PDH相同，后者已被证明包含四个亚基，其分子量与S复合物的分子量非常相似。因此，我们提出S复合物不是蛋白质分泌的主要成分。
