Structure at 2.3 Å resolution of the cytochrome bc1 complex from the yeast Saccharomyces cerevisiae co-crystallized with an antibody Fv fragment
Structure at 2.3 Å resolution of the cytochrome bc1 complex from the yeast Saccharomyces cerevisiae co-crystallized with an antibody Fv fragment
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DOI：
10.1016/s0969-2126(00)00152-0
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发表时间：
2000-06-15
期刊：
影响因子：
5.7
通讯作者：
Michel, H
中科院分区：
文献类型：
--
作者：
Hunte, C;Koepke, J;Michel, H
背景：细胞色素 bc(1) 复合物是呼吸和光合电子传递链能量转换机制的一部分。这种完整的膜蛋白复合物催化从泛醇到细胞色素 c 的电子转移。它将电子转移与质子跨膜的生电易位通过所谓的 Q 循环机制耦合起来。结果：来自酿酒酵母的细胞色素 bc(1) 复合物与结合的抗体 Fv 片段一起结晶。使用多重同晶置换以 2.3 埃分辨率确定结构，并精炼至晶体学 R 因子为 2.2%（无 R = 25.4%）。该复合物以同二聚体形式存在。精制模型的每个“单体”均包含 COR1 亚基的 2178 个氨基酸残基。细胞色素 bc(1) 复合物和 Fv 片段的多肽 V-H 和 V-L、辅因子血红素 b(H)、血红素 b(L)、血红素 c(1)、[2Fe-2S] 簇和 346 个水分子的 QCR2、COB、CVT1、RIP1、QCR6、QCR7、QCR8 和 QCR9。 Fv 片段与 [2Fe-2S] Rieske 蛋白的外在结构域结合，对于晶格的形成至关重要。结论：将膜蛋白结晶为具有特定抗体片段的复合物的方法似乎具有普遍重要性。酵母细胞色素 bc(1) 复合物的结构详细揭示了天然底物辅酶 Q6 和抑制剂 stigmatellin 的结合位点。靠近结合位点的埋藏水分子表明了质子吸收和释放的可能途径。与其他细胞色素 bc(1) 复合物的比较显示了酵母特有的特征。
Background: The cytochrome bc(1) complex is part of the energy conversion machinery of the respiratory and photosynthetic electron transfer chains. This integral membrane protein complex catalyzes electron transfer from ubiquinol to cytochrome c. It couples the electron transfer to the electrogenic translocation of protons across the membrane via a so-called Q cycle mechanism.Results: The cytochrome bc(1) complex from the yeast Saccharomyces cerevisiae was crystallized together with a bound antibody Fv fragment. The structure was determined at 2.3 Angstrom resolution using multiple isomorphous replacement, and refined to a crystallographic R factor of 2.2% (R-free = 25.4%). The complex is present as a homodimer. Each 'monomer' of the refined model includes 2178 amino acid residues of subunits COR1. QCR2, COB, CVT1, RIP1, QCR6, QCR7, QCR8 and QCR9 of the cytochrome bc(1) complex and of the polypeptides V-H and V-L of the Fv fragment, the cofactors heme b(H), heme b(L), heme c(1), the [2Fe-2S] cluster and 346 water molecules. The Fv fragment binds to the extrinsic domain of the [2Fe-2S] Rieske protein and is essential for formation of the crystal lattice.Conclusions: The approach to crystallize membrane proteins as complexes with specific antibody fragments appears to be of general importance. The structure of the yeast cytochrome bc(1) complex reveals in detail the binding sites of the natural substrate coenzyme Q6 and the inhibitor stigmatellin. Buried water molecules close to the binding sites suggest possible pathways for proton uptake and release. A comparison with other cytochrome bc(1) complexes shows features that are specific to yeast.