Departments of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093-0650; Department of Pharmacology, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093-0650.
Department of Pharmacology, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093-0650.
J Biol Chem. 2011 Dec 9;286(49):42555-42565. doi: 10.1074/jbc.M111.286583. Epub 2011 Oct 18.
Determining the structure of the ligand-binding domain of the nicotinic acetylcholine receptor (nAChR) has been a long standing goal in the design of selective drugs useful in implicated diseases for this prevalent receptor family. Acetylcholine-binding proteins have proven to be valuable surrogates with structural similarity and sequence identity to the extracellular domain of the nicotinic receptor, yet these soluble proteins have their unique features and do not serve as exact replicates of the nAChRs of interest. Here we systematically modify the sequence of these proteins toward the homomeric human α7 nAChR. These chimeric proteins exhibit a shift in affinities to reflect α7 binding characteristics yet maintain expression levels and stability conducive for crystallization. We also present a pentameric humanoid nAChR extracellular domain with the structural determination of the α7 nAChR glycosylation site.
确定烟碱型乙酰胆碱受体（nAChR）配体结合域的结构一直是设计用于治疗该普遍受体家族相关疾病的选择性药物的长期目标。乙酰胆碱结合蛋白已被证明是有价值的替代品，它们在结构上与烟碱受体的细胞外结构域相似，并且具有序列同一性，但这些可溶性蛋白具有其独特的特征，不能作为感兴趣的 nAChR 的精确复制品。在这里，我们系统地对这些蛋白质的序列进行修饰，使其接近同源的人α7 nAChR。这些嵌合蛋白的亲和力发生了变化，反映了α7 的结合特性，但仍保持了有利于结晶的表达水平和稳定性。我们还展示了一个五聚体人形烟碱型乙酰胆碱受体细胞外结构域，其中包括α7 nAChR 糖基化位点的结构测定。
