Control of DegP-dependent degradation of c-type cytochromes by heme and the cytochrome c maturation system in Escherichia coli
Control of DegP-dependent degradation of c-type cytochromes by heme and the cytochrome c maturation system in Escherichia coli
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DOI：
10.1128/jb.00656-07
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发表时间：
2007-09-01
影响因子：
3.2
通讯作者：
O'Brian, Mark R.
中科院分区：
文献类型：
--
作者：
Gao, Tao;O'Brian, Mark R.
C型细胞色素部分或全部位于革兰氏阴性细菌的周质中，血红素辅基与蛋白质共价结合。细胞色素c成熟（Ccm）多蛋白系统是运输血红素的周质和其共价键的肽。其他细胞色素和血红蛋白含有非共价结合的血红素，不需要辅助蛋白进行组装。在这里，我们表明，慢生根瘤菌细胞色素c（550）多肽在大肠杆菌中的积累是血红素依赖的，在血红素缺乏的细胞中发现的水平非常低。而周质E.大肠杆菌细胞色素B（562）或胞质Vitrecha血红蛋白（Vh B）的积累与血红素状态无关。细胞色素c（550）的血红素结合半胱氨酸的突变或Ccm的缺乏也导致低载脂蛋白水平。这些水平在degP突变株中恢复，表明脱辅基细胞色素c（550）被周质蛋白酶DegP降解。将细胞色素c血红素结合基序CXXCH引入细胞色素B（562）（c-B（562））中，导致c型细胞色素以Ccm依赖性方式与血红素共价结合。这种变体多肽在血红素缺陷细胞中是稳定的，但在不存在Ccm的情况下被DegP降解。此外，含有周质信号肽和CXXCH基序的Vhb变体不形成C型细胞色素，但积累是Ccm依赖的。数据表明，细胞色素c血红素结合基序是一个不稳定的元素，并通过Ccm的稳定不需要连接的血红素部分的蛋白质。
c-type cytochromes are located partially or completely in the periplasm of gram-negative bacteria, and the heme prosthetic group is covalently bound to the protein. The cytochrome c maturation (Ccm) multiprotein system is required for transport of heme to the periplasm and its covalent linkage to the peptide. Other cytochromes and hemoglobins contain a noncovalently bound heme and do not require accessory proteins for assembly. Here we show that Bradyrhizobium japonicum cytochrome c(550) polypeptide accumulation in Escherichia coli was heme dependent, with very low levels found in heme-deficient cells. However, apoproteins of the periplasmic E. coli cytochrome b(562) or the cytosolic Vitreoscilla hemoglobin (Vhb) accumulated independently of the heme status. Mutation of the heme-binding cysteines of cytochrome c(550) or the absence of Ccm also resulted in a low apoprotein level. These levels were restored in a degP mutant strain, showing that apocytochrome c(550) is degraded by the periplasmic protease DegP. Introduction of the cytochrome c heme-binding motif CXXCH into cytochrome b(562) (c-b(562)) resulted in a c-type cytochrome covalently bound to heme in a Ccm-dependent manner. This variant polypeptide was stable in heme-deficient cells but was degraded by DegP in the absence of Ccm. Furthermore, a Vhb variant containing a periplasmic signal peptide and a CXXCH motif did not form a c-type cytochrome, but accumulation was Ccm dependent nonetheless. The data show that the cytochrome c heme-binding motif is an instability element and that stabilization by Ccm does not require ligation of the heme moiety to the protein.