Castro M J, Kingston I B, Anderson S
Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey 08854, USA.
Anal Biochem. 1995 Apr 10;226(2):225-31. doi: 10.1006/abio.1995.1218.
A simple spectrophotometric assay for the determination of the catalytic efficiency and activity of plasminogen activators is presented. The assay system contains activator, plasminogen, and the chromogenic substrate N-benzoyl-L-arginine-p-nitroanilide (BAPA). Plasmin production is monitored continuously by the hydrolysis of BAPA under non-steady-state, first-order conditions with respect to plasminogen. Apparent catalytic efficiency constants are calculated from the values obtained for the apparent first-order rate constant of activation. The results obtained with the present method were compared with the catalytic efficiency determined through the measurement of kcat and Km, using a different system, under steady-state conditions. Tissue plasminogen activator in the absence and presence of fibrinogen and high-molecular-weight urokinase were used as model activators. Potential applications are discussed.
本文介绍了一种用于测定纤溶酶原激活剂催化效率和活性的简单分光光度法。该检测系统包含激活剂、纤溶酶原和发色底物N-苯甲酰-L-精氨酸对硝基苯胺（BAPA）。在非稳态、纤溶酶原一级条件下，通过BAPA的水解连续监测纤溶酶的产生。根据激活表观一级速率常数获得的值计算表观催化效率常数。将本方法得到的结果与在稳态条件下使用不同系统通过测量kcat和Km确定的催化效率进行了比较。以不存在和存在纤维蛋白原的组织纤溶酶原激活剂以及高分子量尿激酶作为模型激活剂。讨论了潜在的应用。
