Conversion of a c type cytochrome to a b type that spontaneously forms in vitro from apo protein and heme: implications for c type cytochrome biogenesis and folding.

Conversion of a c type cytochrome to a b type that spontaneously forms in vitro from apo protein and heme: implications for c type cytochrome biogenesis and folding.
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c 型细胞色素向 apo 蛋白和血红素在体外自发形成的 b 型转化:对 c 型细胞色素生物发生和折叠的影响。

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发表时间:
2000
影响因子:
11.1
通讯作者:
S. Ferguson
S. Ferguson
中科院分区:
综合性期刊1区
文献类型:
--
作者:
E. Tomlinson;S. Ferguson

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来自嗜热氢杆菌(一种嗜热细菌)的细胞色素 c(552) 在将血红素结合半胱氨酸诱变为丙氨酸并在大肠杆菌细胞质中表达后,已转化为 b 型细胞色素。 b 型变体稳定性较差,盐酸胍解折叠中点的浓度比野生型蛋白低 2 M。还原电位比重组野生型蛋白低75 mV。可以从b型变体中去除血红素,从而产生apo蛋白,根据圆二色光谱,其具有与holo b型蛋白不同的α螺旋含量。后者很容易通过将血红素添加到载脂蛋白中而在体外重新形成。这种重组表明,先前观察到的细胞色素 c(552) 在大肠杆菌细胞质中的组装(具有典型的 I 类细胞色素 c 折叠)是血红素与预折叠多肽结合后自发形成硫醚键的结果。这些观察结果对 c 型细胞色素生物发生的一般问题具有影响。
Cytochrome c(552) from Hydrogenobacter thermophilus, a thermophilic bacterium, has been converted into a b type cytochrome, after mutagenesis of both heme-binding cysteines to alanine and expression in the cytoplasm of Escherichia coli. The b type variant is less stable, with the guanidine hydrochloride unfolding midpoint occurring at a concentration 2 M lower than for the wild-type protein. The reduction potential is 75 mV lower than that of the recombinant wild-type protein. The heme can be removed from the b type variant, thus generating an apo protein that has, according to circular dichroism spectroscopy, an alpha-helical content different from that of the holo b type protein. The latter is readily reformed in vitro by addition of heme to the apo protein. This reforming suggests that previously observed assembly of cytochrome c(552), which has the typical class I cytochrome c fold, in the E. coli cytoplasm is a consequence of spontaneous thioether bond formation after binding of heme to a prefolded polypeptide. These observations have implications for the general problem of c type cytochrome biogenesis.
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