Structurally Based Design of Glucagon Mutants That Inhibit Fibril Formation.
Structurally Based Design of Glucagon Mutants That Inhibit Fibril Formation.
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DOI：
10.1021/acs.biochem.1c00214
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发表时间：
2021-06-29
期刊：
影响因子：
2.9
通讯作者：
Hong M
中科院分区：
文献类型：
--
作者：
Gelenter MD;Dregni AJ;Duan P;Hong M
肽激素胰高血糖素被规定为治疗糖尿病低血糖症的药物化合物。然而，在其高度可溶的酸性pH下，胰高血糖素迅速聚集成无活性和细胞毒性的淀粉样蛋白原纤维。最近确定的高分辨率结构，这些原纤维揭示了各种稳定的分子相互作用。基于这种结构，我们现在已经设计了胰高血糖素的四种精氨酸突变体，其在药物浓度下抵抗凝血数周。S2 R、T29 R双突变体和T29 R单突变体去除野生型原纤维中的氢键相互作用，而Y13 R、A19 R双突变体和Y13 R突变体去除阳离子-π相互作用。1H溶液NMR光谱和UV吸光度数据表明，这些突变体在静止条件下以高达4 mg/ml的浓度在pH 2缓冲液中保持可溶数周。在具有高盐浓度和搅拌的应激条件下，这些突变体的fietize比野生型显著更慢。S2 R、T29 R突变体和T29 R突变体表现出无规卷曲和α-螺旋构象的混合物，而Y13 R突变体完全是无规卷曲。突变位点被选择为不参与与肽的活性结构中的胰高血糖素受体的强相互作用。因此，胰高血糖素的这些精氨酸突变体是治疗低血糖症的有希望的替代化合物。
The peptide hormone glucagon is prescribed as a pharmaceutical compound to treat diabetic hypoglycemia. However, at the acidic pH where it is highly soluble, glucagon rapidly aggregates into inactive and cytotoxic amyloid fibrils. The recently determined high-resolution structure of these fibrils revealed various stabilizing molecular interactions. Based on this structure, we have now designed four arginine mutants of glucagon that resist fibrillization at pharmaceutical concentrations for weeks. An S2R, T29R double mutant and a T29R single mutant remove a hydrogen-bonding interaction in the wild-type fibril, whereas a Y13R, A19R double mutant and a Y13R mutant remove a cation-π interaction. 1H solution NMR spectra and UV absorbance data indicate that these mutants remain soluble in pH 2 buffer under quiescent conditions at concentrations up to 4 mg/ml for weeks. Under stressed conditions with high salt concentrations and agitation, these mutants fibrillize significantly more slowly than the wild-type. The S2R, T29R mutant and the T29R mutant exhibits a mixture of random coil and α-helical conformations while the Y13R mutant is completely random coil. The mutation sites are chosen to be uninvolved in strong interactions with the glucagon receptor in the active structure of the peptide. Therefore, these arginine mutants of glucagon are promising alternative compounds to treat hypoglycemia.
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