Members of the HypC protein family are chaperone-like proteins that play a central role in the maturation of [NiFe]-hydrogenases (Hyd). Escherichia coli has a second copy of HypC, called HybG, and, as a component of the HypDEF maturation scaffold, these proteins help synthesize the NiFe-cofactor and guide the scaffold to its designated hydrogenase large subunit precursor. HypC is required to synthesize active Hyd-1 and Hyd-3, while HybG facilitates Hyd-2 and Hyd-1 synthesis. To identify determinants on HypC that allow it to discriminate against Hyd-2, we made amino acid exchanges in 3 variable regions, termed VR1, VR2, and VR3, of HypC, that make it more similar to HybG. Region VR3 includes a HypC-specific C-terminal α-helical extension, and this proved particularly important in preventing the maturation of Hyd-2 by HypC. Truncation of this extension on HypC increased Hyd-2 activity in the absence of HybG, while retaining maturation of Hyd-3 and Hyd-1. Combining this truncation with amino acid exchanges in VR1 and VR2 of HypC negatively affected the synthesis of active Hyd-1. The C-terminus of E. coli HypC is thus a key determinant in hindering Hyd-2 maturation, while VR1 and VR2 appear more important for Hyd-1 matu-ration.

中文翻译：

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HypC伴侣的扩展C末端α-螺旋限制了[NiFe]-加氢酶成熟过程中Hyp-Scaffold机械对大型亚基前体的识别。

HypC蛋白家族的成员是伴侣蛋白，在[NiFe]-氢化酶（Hyd）的成熟过程中起着关键作用。大肠杆菌具有称为HybG的HypC的第二个副本，并且作为HypDEF成熟支架的组成部分，这些蛋白质有助于合成NiFe辅因子并将支架导向其指定的氢化酶大亚基前体。需要HypC来合成活性Hyd-1和Hyd-3，而HybG可以促进Hyd-2和Hyd-1的合成。为了确定HypC能够区别于Hyd-2的决定因素，我们在3个可变区（称为HypC的VR1，VR2和VR3）中进行了氨基酸交换，这使其与HybG更相似。VR3区域包含一个HypC特异的C端α螺旋延伸，这对于防止HypC成熟的Hyd-2非常重要。在没有HybG的情况下，在HypC上进行此延伸的截短可增加Hyd-2活性，同时保持Hyd-3和Hyd-1的成熟。将此截断与HypC的VR1和VR2中的氨基酸交换结合在一起，会对活性Hyd-1的合成产生负面影响。因此，大肠杆菌HypC的C端是阻碍Hyd-2成熟的关键决定因素，而VR1和VR2似乎对Hyd-1成熟更为重要。