Pb2对核糖核酸酶活性及其结构的影响.docVIP

Pb_2_对核糖核酸酶活性及其结构的影响 化 学 学 报 2003 年第 61 卷 Vol . 61 , 2003 ACTA CHIMICA SINICA 第 1 期 , 117,121 No . 1 , 117,121 2 + Pb对核糖核酸酶活性及其结构的影响 Ξ b a aΞ , a 王 玲 吴 康 王雪峰 洪法水 a b ()苏州大学 生命科学院 图书馆 苏州 215006 陶 冶 ()中国科学院高能物理研究所同步辐射实验室 北京 100039 2 + 2 + 摘要 通过各种光谱学手段研究了 Pb对核糖核酸酶活性的影响及其作用机制. 结果表明低浓度的 Pb可提高酶活性 , 2 + 2 + 高浓度则严重抑制酶活性 ,这是因为在高浓度下 Pb能完全竞争出核糖核酸酶中的 Ca,荧光滴定显示核糖核酸酶可结 2 + 2 + () 合 3 个 Pb. 利用 EXAFS 表征出 Pb已结合到核糖核酸酶主链氨基酸残基上 ,与 N 或 O 发生了配位 ,Pb —N 或 O键长分 2 + 别为 0. 242 nm 和 0. 312 nm ,配位数均为 2. 圆二色谱测试进一步表明高浓度的 Pb结合使核糖核酸酶的二级结构遭到严重 αββ破坏 , 2螺旋含量 、2折叠及2转角大量下降 ,无规则卷曲则明显增加. 2 + 关键词 核糖核酸酶 ,Pb,酶活性 ,配位环境 ,酶结构 2 + Effect of Pbon RNase Activity and Its Structure Ξ , a b a aHONG , Fa2Shui WANG , LingWU , Kang WANG , Xue2Feng a b ( )College of Life Sciences , Library , Suzhou University , Suzhou 215006 TAO , Ye ( )Synchrotron Radiation Laboratory , Institute of High Energy Physics , The Chinese Academy of Sciences , Beijing 100039 2 + (Abstract Activity of RNase from porcine pancreas was enhanced by treatment with Pbat low concentration 0. 17 - 2- 22 + - 22 + μμ) () ×10 ,0. 6 ×10 mol/ Lbut was inhibited by Pbat high concentration over 0. 85 ×10 mol/ L . Pb 2 + at high concentration could competitively displace Cafrom RNase and the fluorescence titration showed that one 2 + 2 + molecule of RNase had three binding sites for Pb, the association constant kfor its low2affinity Pb2binding site sv 6 - 1 2 +() ( ) was 6. 03 ×10mol/ L. The extended X2ray absorption fine structure EXAFSspectrum demonstrated that Pb ()coordinated nitrogen or oxygen atoms of the active site of RNase or other amino acid residue of RNase , Pb —N or O bond length was 0. 242 nm and 0. 312 nm , and coordination number was 2 , respectively. The secondary structure of 2 + αββRNase was greatly damaged by Pbat high concentration resulting in the decrease of 2helix ,2sheet and2turn contents and the increase of random coil contents. 2 + Key words RNase , Pb, enzyme activity , coordination environment , secondary structur