Center for Emerging Infectious Diseases, Gifu University, Yanagido, Japan.
Biophys Chem. 2011 Jun;156(1):24-30. doi: 10.1016/j.bpc.2011.01.009. Epub 2011 Feb 3.
We have used low temperatures (down to -20°C) and high pressures (up to 2000 bar) to populate low-lying excited state conformers of hen lysozyme, and have analyzed their structures site-specifically using (15)N/(1)H two-dimensional HSQC NMR spectroscopy. The resonances of a number of residues were found to be selectively broadened, as the temperature was lowered at a pressure of 2000 bar. The resulting disappearance of cross-peaks includes those of residues in the β-domain of the protein and the cleft between the β- and α-domains, both located close to water-containing cavities. The results indicate that low-lying excited state conformers of hen lysozyme are characterized by slowly fluctuating local conformations around these cavities, attributed to the opportunities for water molecules to penetrate into the cavities. Furthermore, we have found that these water-containing cavities are conserved in similar positions in lysozymes from a range of different biological species, indicating that they are a common evolutionary feature of this family of enzymes.
我们使用低温（低至-20°C）和高压（高达 2000 巴）使母鸡溶菌酶的低能激发态构象稳定，并使用（15）N/（1）H 二维 HSQC NMR 光谱对其结构进行了特异性分析。在 2000 巴的压力下降低温度时，发现许多残基的共振峰被选择性地展宽。由此导致的交叉峰消失包括蛋白质β结构域和β-和α-结构域之间的裂缝中的残基，这些区域都靠近含有水的腔室。结果表明，母鸡溶菌酶的低能激发态构象的特征是这些腔室周围的局部构象缓慢波动，这归因于水分子渗透到腔室中的机会。此外，我们发现这些含水性腔室在来自多种不同生物物种的溶菌酶中处于相似的位置，这表明它们是该酶家族的一个共同进化特征。
