Different residues in periplasmic domains of the CcmC inner membrane protein of Pseudomonas fluorescens ATCC 17400 are critical for cytochrome c biogenesis and pyoverdine-mediated iron uptake
Different residues in periplasmic domains of the CcmC inner membrane protein of Pseudomonas fluorescens ATCC 17400 are critical for cytochrome c biogenesis and pyoverdine-mediated iron uptake
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DOI：
10.1046/j.1365-2958.1998.01085.x
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发表时间：
1998-11-01
影响因子：
3.6
通讯作者：
Cornelis, P
中科院分区：
文献类型：
--
作者：
Gaballa, A;Baysse, C;Cornelis, P
荧光假单胞菌ATCC 17400的内膜蛋白CcmC（CytA）是细胞色素c生物合成所必需的，它在许多细菌和植物线粒体中都有同源物。CcmC缺陷型突变体在绿脓菌荧光素（高亲和力荧光铁载体）的生产和利用中也受到损害。CcmC的拓扑模型，碱性磷酸酶融合的分析的基础上，预测六个跨膜区域与三个周质环。使用定点诱变以评估在所有CcmC同系物中保守的一些周质暴露残基对于细胞色素c生物发生和绿脓菌荧光素生产/利用的重要性。尽管第一周质环中的残基His-61、瓦尔-62和Pro-63以及第三周质环中的Leu-184、His-185和Gln-186保守，但它们同时被Ala替换仅部分影响细胞色素c生物发生和绿脓菌荧光素的产生/利用。第二周质环中残基Trp-115和Gly-116的同时置换显著影响绿脓菌荧光素的产生/利用，但不影响细胞色素c的产生。第二周质环中Asp-127的Ala置换导致细胞色素c的产生减少，在厌氧条件下生长较慢，并且绿脓菌荧光素的产生减少。另一方面，Trp-126中的突变，同样在第二周质环中，完全抑制细胞色素C的产生，而它对绿脓菌荧光素的产生和利用没有影响。这些结果表明，在细胞色素c生物合成和绿脓菌荧光素生产/利用的CcmC的周质结构域中的氨基酸残基的差异参与。
The inner membrane protein CcmC (CytA) of Pseudomonas fluorescens ATCC17400, which has homologues in several bacteria and plant mitochondria, is needed for the biogenesis of cytochrome c. A CcmC-deficient mutant is also compromised in the production and utilization of pyoverdine, the high-affinity fluorescent siderophore. A topological model for CcmC, based on the analysis of alkaline phosphatase fusions, predicts six membrane-spanning regions with three periplasmic loops. Site-directed mutagenesis was used in order to assess the importance of some periplasm-exposed residues, conserved in all CcmC homologues, for cytochrome c biogenesis, and pyoverdine production/utilization. Despite the conservation of the residues His-61, Val-62 and Pro-63 in the first periplasmic loop, and Leu-184, His-185 and Gin-186 in the third periplasmic loop, their simultaneous replacement with Ala only partially affected cytochrome c biogenesis and pyoverdine production/utilization. Simultaneous replacements of residues Trp-115 and Gly-116 in the second periplasmic loop substantially affected pyoverdine production/utilization but not cytochrome c production, An Ala substitution of Asp-127, in the second periplasmic loop, resulted in decreased production of cytochrome c, slower growth in conditions of anaerobiosis and reduced pyoverdine production, On the other hand, a mutation in Trp-126, also in the second periplasmic loop, totally suppressed the production of cytochrome c, whereas it had no effect on the production and utilization of pyoverdine. These results show a differential involvement of amino acid residues in periplasmic domains of CcmC in cytochrome c biogenesis and pyoverdine production/utilization.