Amino acid contribution to protein solubility: Asp, Glu, and Ser contribute more favorably than the other hydrophilic amino acids in RNase Sa.
Amino acid contribution to protein solubility: Asp, Glu, and Ser contribute more favorably than the other hydrophilic amino acids in RNase Sa.
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DOI：
10.1016/j.jmb.2006.10.026
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发表时间：
2007-02
影响因子：
5.6
通讯作者：
Saul R. Trevino;J. Scholtz;C. Pace
中科院分区：
文献类型：
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作者：
Saul R. Trevino;J. Scholtz;C. Pace
蛋白质溶解度差是高分辨率结构研究、蛋白质药物配方和蛋白质生化表征中的常见问题。一个流行的策略，以提高蛋白质的溶解度是使用定点诱变，使疏水到亲水性突变的蛋白质表面。然而，还没有对所有20种氨基酸对蛋白质溶解度的相对贡献进行系统的研究。在这里，在核糖核酸酶（RNase） Sa的完全溶剂暴露位置76处制作了20个变体，以比较每种氨基酸的贡献。还对这些变异进行了稳定性测量，这些变异发生在II型β-转弯的i+1位置。测定了硫酸铵溶液在高净正电荷、低净电荷和高净负电荷条件下的溶解度。令人惊讶的是，即使在亲水性氨基酸中，对蛋白质溶解度也有广泛的贡献。结果表明，天冬氨酸、谷氨酸和丝氨酸的贡献明显优于其他亲水性氨基酸，特别是在高净电荷时。因此，为了增加蛋白质的溶解度，应用天冬氨酸、谷氨酸或丝氨酸代替天冬氨酸、谷氨酰胺或苏氨酸。
Poor protein solubility is a common problem in high-resolution structural studies, formulation of protein pharmaceuticals, and biochemical characterization of proteins. One popular strategy to improve protein solubility is to use site-directed mutagenesis to make hydrophobic to hydrophilic mutations on the protein surface. However, a systematic investigation of the relative contributions of all 20 amino acids to protein solubility has not been done. Here, 20 variants at the completely solvent-exposed position 76 of ribonuclease (RNase) Sa are made to compare the contributions of each amino acid. Stability measurements were also made for these variants, which occur at the i+1 position of a type II β-turn. Solubility measurements in ammonium sulfate solutions were made at high positive net charge, low net charge, and high negative net charge. Surprisingly, there was a wide range of contributions to protein solubility even among the hydrophilic amino acids. The results suggest that aspartic acid, glutamic acid, and serine contribute significantly more favorably than the other hydrophilic amino acids especially at high net charge. Therefore, to increase protein solubility, asparagine, glutamine, or threonine should be replaced with aspartic acid, glutamic acid or serine.