Engineering a minimal G protein to facilitate crystallisation of G protein-coupled receptors in their active conformation.
Engineering a minimal G protein to facilitate crystallisation of G protein-coupled receptors in their active conformation.
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DOI：
10.1093/protein/gzw049
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发表时间：
2016-12
期刊：
影响因子：
--
通讯作者：
Tate CG
中科院分区：
文献类型：
--
作者：
Carpenter B;Tate CG
G蛋白偶联受体（GPCR）响应于细胞外刺激而调节细胞质信号传导，并且是广泛疾病中的重要治疗靶点。GPCR在所有活化状态下的结构测定对于阐明信号转导的精确机制和促进最佳药物设计是重要的。然而，由于其固有的不稳定性，GPCR与细胞质信号传导蛋白如异源三聚体G蛋白和β-抑制蛋白复合物的结晶已被证明具有挑战性。在这里，我们描述了一个最小的G蛋白，迷你GS，这是完全由GTdR结构域的腺苷酸环化酶刺激G蛋白GS的设计。Mini-Gs是一种小的可溶性蛋白，在Gβγ亚基不存在的情况下有效地偶联GPCR。我们利用合理设计诱变技术改造了mini-Gs，使其与去污剂增溶的β1-肾上腺素能受体（β1AR）形成稳定的复合物。迷你G蛋白诱导类似的药理学和结构变化的GPCR作为异源三聚体G蛋白，但消除了许多与这些复合物的结晶，特别是它们的大尺寸，构象动力学和不稳定性在洗涤剂中的问题。因此，它们是新的工具，这将有助于GPCR在其活性构象的生物化学和结构表征。
G protein-coupled receptors (GPCRs) modulate cytoplasmic signalling in response to extracellular stimuli, and are important therapeutic targets in a wide range of diseases. Structure determination of GPCRs in all activation states is important to elucidate the precise mechanism of signal transduction and to facilitate optimal drug design. However, due to their inherent instability, crystallisation of GPCRs in complex with cytoplasmic signalling proteins, such as heterotrimeric G proteins and β-arrestins, has proved challenging. Here, we describe the design of a minimal G protein, mini-Gs, which is composed solely of the GTPase domain from the adenylate cyclase stimulating G protein Gs. Mini-Gs is a small, soluble protein, which efficiently couples GPCRs in the absence of Gβγ subunits. We engineered mini-Gs, using rational design mutagenesis, to form a stable complex with detergent-solubilised β1-adrenergic receptor (β1AR). Mini G proteins induce similar pharmacological and structural changes in GPCRs as heterotrimeric G proteins, but eliminate many of the problems associated with crystallisation of these complexes, specifically their large size, conformational dynamics and instability in detergent. They are therefore novel tools, which will facilitate the biochemical and structural characterisation of GPCRs in their active conformation.
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影响因子：
3.6
作者：
Baker, Jillian G.;Proudman, Richard G. W.;Tate, Christopher G.
通讯作者：
Tate, Christopher G.
DOI：
10.1111/j.1432-1033.1989.tb15068.x
发表时间：
1989-10-01
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
BORNANCIN, F;PFISTER, C;CHABRE, M
通讯作者：
CHABRE, M
影响因子：
64.8
作者：
Huang W;Manglik A;Venkatakrishnan AJ;Laeremans T;Feinberg EN;Sanborn AL;Kato HE;Livingston KE;Thorsen TS;Kling RC;Granier S;Gmeiner P;Husbands SM;Traynor JR;Weis WI;Steyaert J;Dror RO;Kobilka BK
通讯作者：
Kobilka BK
影响因子：
64.8
作者：
Flock T;Ravarani CNJ;Sun D;Venkatakrishnan AJ;Kayikci M;Tate CG;Veprintsev DB;Babu MM
通讯作者：
Babu MM
影响因子：
64.8
作者：
Lebon, Guillaume;Warne, Tony;Edwards, Patricia C.;Bennett, Kirstie;Langmead, Christopher J.;Leslie, Andrew G. W.;Tate, Christopher G.
通讯作者：
Tate, Christopher G.