Investigation of the regiospecificity and stereospecificity of proton transfer in the yeast inorganic pyrophosphatase catalyzed reaction.
Investigation of the regiospecificity and stereospecificity of proton transfer in the yeast inorganic pyrophosphatase catalyzed reaction.
复制标题
酵母无机焦磷酸酶催化反应中质子转移的区域特异性和立体特异性的研究。
DOI：
10.1021/bi00364a035
复制
发表时间：
1986
期刊：
影响因子：
2.9
通讯作者：
Dunaway-Mariano,D
中科院分区：
文献类型：
--
作者：
Lin,I;Knight,WB;Hsueh,A;Dunaway-Mariano,D
根据Cooperaran等的改良方法（Bond，1979）纯化PPase。（1973年）。这些实验中使用的酶在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳凝胶（7.5%丙烯酰胺）上迁移为单一条带，活性为690 µ P·min-1（pH 7.5时蛋白质的mg/min）。所有PPase浓度均以活性位点浓度报告。根据Cornelius等人的方法制备了P1，P2-双齿Co（N-H3）4PP和P1，P2-双齿Co（NH3）4PNP。（1977）和Haromy et al.（1983）。在IBM WP 200 SY（在81.02Hz下操作）NMR光谱仪上测量31 P NMR光谱。化学位移以ppm为单位报告，低场（+）或高场（-）为0.1 Md 3 pg 4外标物。
Materials and MethodsPPase was purified according to the modified method (Bond, 1979) of Cooperaran et al.(1973). The enzyme used in these experiments migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels (7.5% acrylamide) and had an activity of 690 µ of P¡ min" 1 (mg of protein)'1 at pH 7.5. All PPase concentrations are reported in terms of active site concentrations. P ‘, P2-Bidentate Co (N-H3) 4PP and P1, P2-bidentate Co (NH3) 4PNP were prepared according to the methods of Cornelius et al.(1977) and Haromy et al.(1983), respectively. 31P NMR spectra were measured on an IBM WP200SY (operating at 81.02 Hz) NMR spectrometer. Chemical shifts are reported in ppm downfield (+) or upfield (-) from a 0.1 Md3pg4 external standard.