标题：A Simple and Efficient Method for Preparation of Monomethoxypolyethylene Glycol Activated with p-Nitrophenylchloroformate and Its Application to Modification of L-Asparaginase
期刊名称：Journal of Clinical Biochemistry and Nutrition
印刷版ISSN：0912-0009
电子版ISSN：1880-5086
出版年度：1996
卷号：21
期号：2
页码：101-111
DOI：10.3164/jcbn.21.101
出版社：The Society for Free Radical Research Japan
摘要：An improved, simple and efficient method for preparation of monomethoxypolyethylene glycol (PEG) activated with p -nitrophenylchloroformate (PNP-PEG) and its use as a potent modifier of protein under mild conditions are described. Modification of bovine serum albumin with PNP-PEG was compared with that done with PEG activated with N, N′-carbonyldiimidazole or cyanuric chloride. The reaction of PEG, activated with either p -nitrophenylchloroformate or cyanuric chloride, with bovine serum albumin at 4°C reached a plateau within 1h, whereas protein modification using PEG activated with N, N′-carbonyldiimidazole was rather slow and gave a low yield. The remaining activity of L-asparaginase modified with PNP-PEG was much higher than that of the enzyme modified to the same degree with PEG activated with cyanuric chloride. At a 20 molar excess of PNP-PEG having a molecular weight of 5, 000, 55% of the free amino acid groups were modified at 4°C for 2h, and the modified enzyme still had 33% residual enzyme activity. Immunochemical studies showed that the highly modified enzyme (67% modification with 18% residual enzyme activity) had lost its immunogenicity and had become much less sensitive to protease digestion.
