Luchter-Wasylewska E, Dulińska J, Ostrowski W S, Torchilin V P, Trubetskoy V S
Institute of Medical Biochemistry, Medical Academy, Kraków, Poland.
Biotechnol Appl Biochem. 1991 Feb;13(1):36-47.
Human prostatic acid phosphatase (PAP) (EC 3.1.3.2) was covalently linked to chondroitin sulfate A from whale cartilage. In order to bind the protein amino groups with the preactivated carboxyl groups of chondroitin sulfate, 1-ethyl-3-(3'-dimethylaminepropyl)carbodiimide and N-hydroxysulfosuccinimide were used as coupling agents. The product was soluble and enzymatically active. The activity was on average 25% higher than that of the free enzyme. The product was heterogeneous in respect to charge and Mr (50-1500) kDa, as determined by chromatography on Sephacryl S 300 and polyacrylamide gel electrophoresis. The resulting polymers contained covalently bound chondroitin sulfate, as shown by the biotin-avidin test. The modified enzyme is more resistant against various denaturing agents, e.g., urea, ethanol, and heat. Thus covalent modification of PAP by cross-linking to chondroitin sulfate could be the preferred method for stabilization of its biological activity.
人前列腺酸性磷酸酶（PAP）（EC 3.1.3.2）与来自鲸软骨的硫酸软骨素A共价连接。为了使蛋白质氨基与硫酸软骨素的预活化羧基结合，使用1-乙基-3-（3'-二甲氨基丙基）碳二亚胺和N-羟基磺基琥珀酰亚胺作为偶联剂。产物可溶且具有酶活性。其活性平均比游离酶高25%。通过Sephacryl S 300色谱和聚丙烯酰胺凝胶电泳测定，产物在电荷和分子量（50 - 1500）kDa方面具有异质性。生物素 - 抗生物素蛋白试验表明，所得聚合物含有共价结合的硫酸软骨素。修饰后的酶对各种变性剂（如尿素、乙醇和热）具有更强的抗性。因此，通过与硫酸软骨素交联对PAP进行共价修饰可能是稳定其生物活性的首选方法。
