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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >A defined protein-detergent-lipid complex for crystallization of integral membrane proteins: The cytochrome b6f complex of oxygenic photosynthesis.
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A defined protein-detergent-lipid complex for crystallization of integral membrane proteins: The cytochrome b6f complex of oxygenic photosynthesis.

机译：用于结晶完整膜蛋白的明确的蛋白质-洗涤剂-脂质复合物：氧光合作用的细胞色素b6f复合物。

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摘要

The paucity of integral membrane protein structures creates a major bioinformatics gap, whose origin is the difficulty of crystallizing these detergent-solubilized proteins. The problem is particularly formidable for hetero-oligomeric integral membrane proteins, where crystallization is impeded by the heterogeneity and instability of the protein subunits and the small lateral pressure imposed by the detergent micelle envelope that surrounds the hydrophobic domain. In studies of the hetero (eight subunit)-dimeric 220,000 molecular weight cytochrome b(6)f complex, derived from the thermophilic cyanobacterium, Mastigocladus laminosus, crystals of the complex in an intact state could not be obtained from highly purified delipidated complex despite exhaustive screening. Crystals of proteolyzed complex could be obtained that grew very slowly and diffracted poorly. Addition to the purified lipid-depleted complex of a small amount of synthetic nonnative lipid, dioleolyl-phosphatidylcholine, resulted in a dramatic improvement in crystallization efficiency. Large crystals of the intact complex grew overnight, whose diffraction parameters are as follows: 94% complete at 3.40 A spacing; R(merge) = 8.8% (38.5%), space group, P6(1)22; and unit cell parameters, a = b = 156.3 A, c = 364.0 A, alpha = beta = 90(o), gamma = 120(o). It is proposed that the methodology of augmentation of a well-defined lipid-depleted integral membrane protein complex with synthetic nonnative lipid, which can provide conformational stability to the protein complex, may be of general use in the crystallization of integral membrane proteins.
机译：缺乏完整的膜蛋白结构造成了主要的生物信息学鸿沟，其起源是难以使这些洗涤剂溶解的蛋白结晶。对于异寡聚体整体膜蛋白而言，这个问题尤为严峻，在这种情况下，蛋白质亚基的异质性和不稳定性以及包围疏水域的去污剂胶束包膜所施加的较小的侧向压力阻碍了结晶。在对嗜热蓝细菌Mastigocladus laminosus衍生的异质（八个亚基）-二聚体分子量为220,000的细胞色素b（6）f复合物的研究中，尽管详尽无遗，仍无法从高度纯化的脱脂复合物中获得完整状态的复合物晶体。筛选。可以得到蛋白水解的复合物晶体，其生长非常缓慢并且衍射差。将少量合成的非天然脂质，二油酰基-磷脂酰胆碱加到纯化的贫脂质复合物中，可显着提高结晶效率。完整复合物的大晶体生长过夜，其衍射参数如下：94％的分子以3.40 A的间隔完成； R（合并）= 8.8％（38.5％），空间群，P6（1）22;和晶胞参数，a = b = 156.3 A，c = 364.0 A，alpha = beta = 90（o），gamma = 120（o）。有人提出，可以为蛋白质复合物提供构象稳定性的，用合成的非天然脂质增加定义明确的脂质耗尽的整体膜蛋白复合物的方法，可能是在整体膜蛋白的结晶中普遍使用的方法。

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