Central modules of the vaccinia virus complement control protein are not in extensive contact
Central modules of the vaccinia virus complement control protein are not in extensive contact
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DOI：
10.1042/0264-6021:3440167
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发表时间：
1999-11-15
影响因子：
4.1
通讯作者：
Barlow, PN
中科院分区：
文献类型：
--
作者：
Kirkitadze, MD;Henderson, C;Barlow, PN
28.6 kDa的牛痘病毒补体控制蛋白（VCP）是补体系统的抑制剂，具有治疗潜力。它由四个结构域或模块组成，是补体受体1（CR 1）和其他哺乳动物补体激活调节因子的同源物。这些蛋白质中结构-功能关系的一个关键方面是分子内模块-模块相互作用的程度，因为这些决定了分子的整体形状和灵活性。在毕赤酵母中高效表达了包含VCP模块2和3的蛋白片段（VCP类似于2，3）。超离心表明，类似于2，3的VCP是高度不对称的，轴比为5.3：1，这与两个模块的端对端排列一致。NMR光谱、差示扫描量热法、CD和固有色氨酸荧光用于监测类似于2，3的VCP的解折叠。在一系列的温度和浓度的氯化胍进行的实验表明，模块2展开在温和的条件下比，并独立于，模块3。模块2的展开与模块3的酰胺N-15和H-1化学位移的广泛变化无关，这意味着模块不形成广泛的模块间界面。将在这项工作中获得的类似于2，3的VCP的结果与在CRI模块15-17的研究中获得的结果进行比较[Kirkitadze，Krych，Uhrin，德莱登，Smith，库珀，Wang，Hauhart，Atkinson和Barlow（1999）Biochemistry 38，7019-7031]。
The 28.6 kDa vaccinia virus complement control protein (VCP) is an inhibitor of the complement system and has therapeutic potential. It is composed of four domains or modules and is a homologue of complement receptor 1 (CR1) and other mammalian regulators of complement activation. A key aspect to structure-function relationships in these proteins is the extent of intramolecular module-module interactions, since these dictate the overall shape and flexibility of the molecules. A protein fragment (VCP similar to 2,3) encompassing modules 2 and 3 of VCP was over-expressed in Pichia pastoris. Ultracentrifugation showed that VCP similar to 2,3 is highly asymmetric with an axial ratio of 5.3: 1, which is consistent with an end-to-end arrangement of the two modules. NMR spectroscopy, differential scanning calorimetry, CD and intrinsic tryptophan fluorescence were used to monitor unfolding of VCP similar to 2,3. Experiments performed over a range of temperatures and concentrations of guanidinium chloride revealed that module 2 unfolds under milder conditions than, and independently of, module 3. Unfolding of module 2 is not associated with extensive changes in amide N-15 and H-1 chemical shifts of module 3, implying that the modules do not form an extensive intermodular interface. Results obtained in this work for VCP similar to 2,3 are compared with those obtained in a study of CRI modules 15-17 [Kirkitadze, Krych, Uhrin, Dryden, Smith, Cooper, Wang, Hauhart, Atkinson and Barlow (1999) Biochemistry 38, 7019-7031].