Mutagenic and thermodynamic analyses of residual structure in the alpha subunit of tryptophan synthase.
Mutagenic and thermodynamic analyses of residual structure in the alpha subunit of tryptophan synthase.
复制标题
色氨酸合酶α亚基残余结构的诱变和热力学分析。
DOI：
10.1021/bi951726o
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Matthews,CR
中科院分区：
文献类型：
--
作者：
Saab-Rincon,G;Gualfetti,PJ;Matthews,CR
来自大肠杆菌的色氨酸合酶的α亚基先前已显示在5 M尿素下含有残余结构，在该条件下二级结构完全破坏并且酪氨酸残基暴露于溶剂[Saab-Rincón，G.，福罗布角L.，&马修斯，C. R.（1993）Biochemistry32，13981 - 13990]。可以通过组氨酸92的一维NMR光谱研究来监测残留结构，组氨酸92的Cε质子对该形式与未折叠蛋白质之间的缓慢交换敏感。中间体和未折叠形式之间的协同尿素诱导的未折叠转变的温度依赖性表明，该过程在25 °C下涉及焓变和熵变的负值。组氨酸92附近的几个非极性侧链的替代对展开过渡曲线的斜率和中点的影响表明，这些侧链参与残余结构。然而，跨越组氨酸92和突变残基的15个残基的肽不足以定义该结构。这些结果表明，α亚基中的残留结构通过疏水效应稳定，可能涉及与组氨酸92序列较远的侧链。
The α subunit of tryptophan synthase fromEscherichia colihas been previously shown to contain residual structure at 5 M urea, conditions where the secondary structure is entirely disrupted and the tyrosine residues are exposed to solvent [Saab-Rincón, G., Froebe, C. L., & Matthews, C. R. (1993)Biochemistry32, 13981−13990]. The residual structure can be monitored by one-dimensional NMR spectroscopy studies of histidine 92 whose Cε proton is sensitive to the slow exchange between this form and the unfolded protein. The temperature dependence of the cooperative urea-induced unfolding transition between intermediate and unfolded forms demonstrates that this process involves negative values for both the enthalpy and entropy changes at 25 °C. The effects of replacements of several nonpolar side chains adjacent to histidine 92 on the slopes and midpoints of the unfolding transition curve show that these side chains participate in the residual structure. A 15-residue peptide spanning histidine 92 and the mutated residues, however, is not sufficient to define this structure. These results demonstrate that the residual structure in the α subunit is stabilized by the hydrophobic effect and may involve side chains which are distant in sequence to histidine 92.