Yagi Daichi, Yamada Taro, Kurihara Kazuo, Ohnishi Yuki, Yamashita Masahiro, Tamada Taro, Tanaka Ichiro, Kuroki Ryota, Niimura Nobuo
Graduate School of Science and Engineering, Ibaraki University, Hitachi, Japan.
Acta Crystallogr D Biol Crystallogr. 2009 Sep;65(Pt 9):892-9. doi: 10.1107/S0907444909018885. Epub 2009 Aug 6.
A neutron crystallographic analysis of phosphate-free bovine pancreatic RNase A has been carried out at 1.7 A resolution using the BIX-4 single-crystal diffractometer at the JRR-3 reactor of the Japan Atomic Energy Agency. The high-resolution structural model allowed us to determine that His12 acts mainly as a general base in the catalytic process of RNase A. Numerous other distinctive structural features such as the hydrogen positions of methyl groups, hydroxyl groups, prolines, asparagines and glutamines were also determined at 1.7 A resolution. The protonation and deprotonation states of all of the charged amino-acid residues allowed us to provide a definitive description of the hydrogen-bonding network around the active site and the H atoms of the key His48 residue. Differences in hydrogen-bond strengths for the alpha-helices and beta-sheets were inferred from determination of the hydrogen-bond lengths and the H/D-exchange ratios of the backbone amide H atoms. The correlation between the B factors and hydrogen-bond lengths of the hydration water molecules was also determined.
利用日本原子能机构JRR - 3反应堆的BIX - 4单晶衍射仪，在1.7 Å分辨率下对无磷酸牛胰核糖核酸酶A进行了中子晶体学分析。高分辨率结构模型使我们能够确定，His12在核糖核酸酶A的催化过程中主要作为一般碱起作用。在1.7 Å分辨率下还确定了许多其他独特的结构特征，如甲基、羟基、脯氨酸、天冬酰胺和谷氨酰胺的氢位置。所有带电荷氨基酸残基的质子化和去质子化状态使我们能够对活性位点周围的氢键网络以及关键His48残基的H原子提供明确描述。通过测定氢键长度和主链酰胺H原子的H/D交换率，推断出α螺旋和β折叠的氢键强度差异。还确定了水合水分子的B因子与氢键长度之间的相关性。
