Overlapping specificities of the mitochondrial cytochrome c and c1 heme lyases
Overlapping specificities of the mitochondrial cytochrome c and c1 heme lyases
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DOI：
10.1074/jbc.m308881200
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发表时间：
2003-12-12
影响因子：
4.8
通讯作者：
Hamel, PP
中科院分区：
文献类型：
--
作者：
Bernard, DG;Gabilly, ST;Hamel, PP
血红素附着到真菌线粒体细胞色素c和c（1）的脱辅基型需要细胞色素c和c（1）血红素裂解酶（CCHL和CC 1HL）的活性，这是具有不同底物特异性的酶。然而，在高等真核生物中存在单一的血红素裂解酶表明了更广泛的底物特异性。在这里，我们证明了酵母CCHL对非同源底物脱辅基细胞色素c（1）是有活性的，即CCHL促进低水平的脱辅基细胞色素c（1）转化为它的全形在CC 1HL的情况下。此外，单一的人血红素裂解酶还显示出更广泛的细胞色素特异性，这从其替代酵母CCHL和CC 1HL的能力中显而易见。分离出CC 1HL缺失的多拷贝和遗传抑制子，其分析显示CCHL对细胞色素c（1）的活性可通过以下方式增强：1）降低同源底物脱辅基细胞色素c的丰度，2）增加CCHL的积累，3）通过CCHL或细胞色素c（1）中的点突变改变底物-酶相互作用，或4）过表达Cyc 2 p，一种以前只知道是线粒体生物发生因子的蛋白质。基于Cyc 2 p与CCHL的功能相互作用和蛋白质中存在假定的FAD结合位点，我们假设Cyc 2 p控制血红素裂解酶反应的氧化还原化学。
Heme attachment to the apoforms of fungal mitochondrial cytochrome c and c(1) requires the activity of cytochrome c and c(1) heme lyases (CCHL and CC1HL), which are enzymes with distinct substrate specificity. However, the presence of a single heme lyase in higher eukaryotes is suggestive of broader substrate specificity. Here, we demonstrate that yeast CCHL is active toward the non-cognate substrate apocytochrome c(1), i.e. CCHL promotes low levels of apocytochrome c(1) conversion to its holoform in the absence of CC1HL. Moreover, that the single human heme lyase also displays a broader cytochrome specificity is evident from its ability to substitute for both yeast CCHL and CC1HL. Multicopy and genetic suppressors of the absence of CC1HL were isolated and their analysis revealed that the activity of CCHL toward cytochrome c(1) can be enhanced by: 1) reducing the abundance of the cognate substrate apocytochrome c, 2) increasing the accumulation of CCHL, 3) modifying the substrate-enzyme interaction through point mutations in CCHL or cytochrome c(1), or 4) overexpressing Cyc2p, a protein known previously only as a mitochondrial biogenesis factor. Based on the functional interaction of Cyc2p with CCHL and the presence of a putative FAD-binding site in the protein, we hypothesize that Cyc2p controls the redox chemistry of the heme lyase reaction.