利用紫外光谱研究大肠杆菌核糖核酸酶H和限制性内切核酸酶对核酸的切割作用。
Use of UV spectroscopy for the study of nucleic acid cleavage by E. coli RNase H and restriction endonucleases.
作者信息
Petrauskene O V, Krynetskaya N F, Tashlitsky V N, Belkov V M, Kubareva E A, Gromova E S, Guschlbauer W, Shabarova Z A
机构信息
Department of Chemistry, Moscow State University, Russia.
出版信息
Biochem Mol Biol Int. 1995 Dec;37(6):1127-35.
A one-step spectrophotometric method for monitoring of nucleic acid cleavage by ribonuclease H from E. coli and type II restriction endonucleases has been proposed. It is based on recording of the increase in the UV absorbance at 260 nm during the course of enzymatic reaction. Duplexes stable under the reaction conditions were chosen as substrates for the enzymes being studied. In order to obtain duplex dissociation following their cleavage by the enzyme appreciate temperature conditions were selected. The spectrophotometric method may be applied for rapid testing of the nuclease activity in protein preparations as well as for precise quantitative analysis of nucleic acid degradation by enzymes. This method may be successfully employed in kinetic studies of nucleic acid-protein interactions.
提出了一种用于监测大肠杆菌核糖核酸酶H和II型限制性核酸内切酶切割核酸的一步分光光度法。它基于在酶促反应过程中记录260nm处紫外吸光度的增加。选择在反应条件下稳定的双链体作为所研究酶的底物。为了在酶切割后获得双链体解离，选择了合适的温度条件。该分光光度法可用于快速检测蛋白质制剂中的核酸酶活性，以及对酶促核酸降解进行精确的定量分析。该方法可成功应用于核酸-蛋白质相互作用的动力学研究。
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