THERMODYNAMIC ANALYSIS OF THE FOLDING OF THE STREPTOCOCCAL PROTEIN-G IGG-BINDING DOMAINS B1 AND B2 - WHY SMALL PROTEINS TEND TO HAVE HIGH DENATURATION TEMPERATURES
THERMODYNAMIC ANALYSIS OF THE FOLDING OF THE STREPTOCOCCAL PROTEIN-G IGG-BINDING DOMAINS B1 AND B2 - WHY SMALL PROTEINS TEND TO HAVE HIGH DENATURATION TEMPERATURES
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DOI：
10.1021/bi00129a007
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发表时间：
1992-04-14
期刊：
影响因子：
2.9
通讯作者：
BRYAN, P
中科院分区：
文献类型：
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作者：
ALEXANDER, P;FAHNESTOCK, S;BRYAN, P
我们克隆、表达并鉴定了链球菌蛋白g的igg结合结构域的两种自然变异。该结构域是一个由56个氨基酸组成的稳定的协同折叠单元，它保持着独特的折叠结构，没有二硫交联或紧密的配体结合。我们研究了这个结构域的两个版本的展开反应的热力学，称为B1和B2，它们的区别是6个氨基酸。通过差示扫描量热法测定，在pH值5.4时，它们的变性温度分别为87.5℃和79.4℃。展开反应的热力学状态函数（DELTA-G, DELTA-H， DELTA-S和DELTA-C(p)）已经确定，并揭示了一些非常小的蛋白质行为的有趣见解。首先，尽管B1结构域具有接近90℃的热变性点，但在生理相关温度下并不异常稳定（37℃时δ - g = 25 kJ/mol）。这种行为的发生是因为稳定性曲线（DELTA-G与温度）平坦而浅，这是由于展开时的DELTA-S和DELTA-C(p)较小。与这一点相关的是第二个观察结果，即由于溶剂条件的突变或变化而导致的b畴展开自由能的微小变化会导致热变性温度的大变化。第三，疏水与非疏水力（每个氨基酸残基）对b结构域折叠总自由能的大小和相对贡献与其他大得多的球状蛋白质非常典型。
We have cloned, expressed, and characterized two naturally occurring variations of the IgG-binding domain of streptococcal protein G. The domain is a stable cooperative folding unit of 56 amino acids, which maintains a unique folded structure without disulfide cross-links or tight ligand binding. We have studied the thermodynamics of the unfolding reaction for the two versions of this domain, designated B1 and B2, which differ by six amino acids. They have denaturation temperatures of 87.5-degrees-C and 79.4-degrees-C, respectively at pH 5.4, as determined by differential scanning calorimetry. Thermodynamic state functions for the unfolding reaction (DELTA-G, DELTA-H, DELTA-S, and DELTA-C(p)) have been determined and reveal several interesting insights into the behavior of very small proteins. First, though the B1 domain has a heat denaturation point close to 90-degrees-C, it is not unusually stable at physiologically relevant temperatures (DELTA-G = 25 kJ/mol at 37-degrees-C). This behavior occurs because the stability profile (DELTA-G vs temperature) is flat and shallow due to the small DELTA-S and DELTA-C(p) for unfolding. Related to this point is the second observation that small changes in the free energy of unfolding of the B-domain due to mutation or change in solvent conditions lead to large shifts in the heat denaturation temperature. Third, the magnitude and relative contributions of hydrophobic vs nonhydrophobic forces (per amino acid residue) to the total free energy of folding of the B-domain are remarkably typical of other globular proteins of much larger size.