A cytochrome c biogenesis gene involved in pyoverdine production in Pseudomonas fluorescens ATCC 17400
A cytochrome c biogenesis gene involved in pyoverdine production in Pseudomonas fluorescens ATCC 17400
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DOI：
10.1046/j.1365-2958.1996.391399.x
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发表时间：
1996-08-01
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;Koedam, N;Cornelis, P
荧光假单胞菌atcc 17400在铁限制条件下产生吡哆醇。Tn5突变体2G11产量较低，在乙二胺-二(邻羟基苯乙酸)(EDDHA)或锌引起的铁限制下不能生长。该突变体还含有一个9.6kb的HindIII-BamHI DNA片段，该片段包含8个连续的开放阅读框(ORF、CyTA到CytH)。该多顺反子基因簇可能编码的蛋白质不仅在氨基酸序列上，而且在总亲水性指数上都与来自胶囊红杆菌和日本慢生根瘤菌等细胞色素c生物发生基因的产物相似。通过TnPhoA突变和定点基因替换，发现前三个ORF(CyTA到CytC)对细胞色素c的产生是必需的，而只有CyTA的产物是正常产生吡喃佛定所必需的。证实了在CyTA蛋白(WGSWWVWD)中存在一个可能的血红素结合位点。通过对构建的PhoA融合的分析，发现了该基序的周质位置。CyTA基因能够恢复细胞色素c和吡喃二酚的产生，这表明该基因参与了荧光假单胞菌的血红素和吡喃二磷酸的运输。
Pseudomonas fluorescens ATCC 17400 produces pyoverdine under iron-limiting conditions. A Tn5 mutant, 2G11, produced lower amounts of different pyoverdine forms and was unable to grow under iron limitation caused by ethylenediamine-di(o-hydroxy-phenylacetic acid) (EDDHA) or zinc. This mutant was complemented by a 9.6 kb HindIII-BamHI DNA fragment that contained eight contiguous open reading frames (ORFs cytA to cytH). The proteins possibly encoded by this polycistronic gene cluster were all similar to the products of cytochrome c biogenesis genes from, amongst others, Rhodobacter capsulatus and Brady-rhizobium japonicum, not only in terms of amino acid sequence, but also in the overall hydropathy index of these proteins. By TnphoA mutagenesis and site-specific gene replacement it was found that the first three ORFs (cytA to cytC) were essential for cytochrome c production while only the product of cytA was needed for normal pyoverdine production. The presence of a putative haem-binding site in the CytA protein (WGSWWVWD) was confirmed. From analysis of a constructed phoA fusion, a periplasmic location was found for this motif. The ability of the cytA gene to restore both cytochrome c and pyoverdine production suggests the involvement of this particular gene both in haem and in pyoverdine transport in P. fluorescens.