Lang S E, Jenney F E, Daldal F
Department of Biology, Plant Science Institute, University of Pennsylvania, Philadelphia 19104-6018, USA.
J Bacteriol. 1996 Sep;178(17):5279-90. doi: 10.1128/jb.178.17.5279-5290.1996.
While searching for components of the soluble electron carrier (cytochrome c2)-independent photosynthetic (Ps) growth pathway in Rhodobacter capsulatus, a Ps- mutant (FJM13) was isolated from a Ps+ cytochrome c2-strain. This mutant could be complemented to Ps+ growth by cycA encoding the soluble cytochrome c2 but was unable to produce several c-type cytochromes. Only cytochrome c1 of the cytochrome bc1 complex was present in FJM13 cells grown on enriched medium, while cells grown on minimal medium contained at various levels all c-type cytochromes, including the membrane-bound electron carrier cytochrome cy. Complementation of FJM13 by a chromosomal library lacking cycA yielded a DNA fragment which also complemented a previously described Ps- mutant, MT113, known to lack all c-type cytochromes. Deletion and DNA sequence analyses revealed an open reading frame homologous to cycH, involved in cytochrome c biogenesis. The cycH gene product (CycH) is predicted to be a bipartite protein with membrane-associated amino-terminal (CycH1) and periplasmic carboxyl-terminal (CycH2) subdomains. Mutations eliminating CyCH drastically decrease the production or all known c-type cytochromes. However, mutations truncating only its CycH2 subdomain always produce cytochrome c1 and affect the presence of other cytochromes to different degrees in a growth medium-dependent manner. Thus, the subdomain CycH1 is sufficient for the proper maturation of cytochrome c1 which is the only known c-type cytochrome anchored to the cytoplasmic membrane by its carboxyl terminus, while CycH2 is required for efficient biogenesis of other c-type cytochromes. These findings demonstrate that the two subdomains of CycH play different roles in the biogenesis of topologically distinct c-type cytochromes and reconcile the apparently conflicting data previously obtained for other species.
在寻找荚膜红细菌中不依赖可溶性电子载体（细胞色素c2）的光合（Ps）生长途径的组成成分时，从一株Ps⁺细胞色素c2⁻菌株中分离出了一个Ps⁻突变体（FJM13）。该突变体可通过编码可溶性细胞色素c2的cycA互补为Ps⁺生长，但无法产生几种c型细胞色素。在富集培养基上生长的FJM13细胞中仅存在细胞色素bc1复合体的细胞色素c1，而在基本培养基上生长的细胞则含有不同水平的所有c型细胞色素，包括膜结合电子载体细胞色素cy。用缺乏cycA的染色体文库对FJM13进行互补，得到了一个DNA片段，该片段也能互补先前描述的已知缺乏所有c型细胞色素的Ps⁻突变体MT113。缺失和DNA序列分析揭示了一个与参与细胞色素c生物合成的cycH同源的开放阅读框。cycH基因产物（CycH）预计是一种具有膜相关氨基末端（CycH1）和周质羧基末端（CycH2）亚结构域的双功能蛋白。消除CycH的突变会大幅降低所有已知c型细胞色素的产生。然而，仅截断其CycH2亚结构域的突变总是会产生细胞色素c1，并以生长培养基依赖的方式不同程度地影响其他细胞色素的存在。因此，亚结构域CycH1足以使细胞色素c1正确成熟，细胞色素c1是唯一已知的通过其羧基末端锚定在细胞质膜上的c型细胞色素，而CycH2是其他c型细胞色素高效生物合成所必需的。这些发现表明，CycH的两个亚结构域在拓扑结构不同的c型细胞色素的生物合成中发挥着不同的作用，并协调了先前在其他物种中获得的明显相互矛盾的数据。
