A cavity-filling mutation at a hydrophobic cavity is a useful method for increasing protein stability. This method, however, sometimes destabilizes the protein because of the accompanying structural changes by the steric hindrance around the cavity. Thus, detailed knowledge of unfavorable structural changes is important for a comprehensive understanding of the cavity-filling mutation. In the present study, by employing the cavity-filling mutant of Escherichia coli RNase HI as a case study, the structural change induced by the substitution of Phe for Ala52 (Ala52Phe) was analyzed in detail using Raman spectroscopy. In previous studies, the thermodynamic result apparently indicated a small decrease in ΔG (destabilization) by the mutation. In the present study, Raman differential spectra show a clear structural difference between wild-type E. coli RNase HI and Ala52Phe. Consequently, the direct signature of the conformational strains around the protein cavity is readily acquired, leading to further understanding of the trade-off relationship between the cavity-filling and incidental steric hindrance.

中文翻译：

---

大肠杆菌RNase HI腔填充不稳定突变蛋白中Steric菌株的光谱签名。

疏水腔处的腔填充突变是增加蛋白质稳定性的有用方法。然而，由于伴随腔周围空间位阻的伴随结构变化，该方法有时使蛋白质不稳定。因此，对不利的结构变化的详细了解对于全面了解腔填充突变很重要。在本研究中，以大肠杆菌RNase HI的空腔填充突变体为例，通过拉曼光谱法详细分析了用Phe替代Ala52（Ala52Phe）引起的结构变化。在以前的研究中，热力学结果显然表明该突变使ΔG（稳定化）略有下降。在本研究中，拉曼光谱显示野生型E之间存在明显的结构差异。大肠杆菌RNase HI和Ala52Phe。因此，很容易获得围绕蛋白质腔的构象菌株的直接特征，从而进一步了解了腔填充与偶然空间位阻之间的权衡关系。