Preparation and characterization of a bovine serum albumin-catalase conjugate

Chinese Academy of Sciences
Biotechnology Letters (Impact Factor: 1.59). 05/2002; 24(12):1027-1030. DOI: 10.1023/A:1015641416286


A bovine serum albumin-catalase conjugate was prepared using a heterobifunctional reagent, N-succinimidyl 3-(2-pyridyldithio)propionate. The purified conjugate had an M
r of 380 kDa. The half-life time of the conjugate was 6, 69 and 65 min after heat treatment, urea treatment and trypsin hydrolysis, respectively, whereas that of native catalase was 3, 22, 28 min, respectively.

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    ABSTRACT: Ribonuclease A (RNase A) is a therapeutic enzyme with cytotoxic action against tumor cells. Its clinical application is limited by the short half-life and insufficient stability. Conjugation of albumin can overcome the limitation, whereas dramatically decrease the enzymatic activity of RNase A. Here, three strategies were proposed to prepare the RNase A–bovine serum albumin (BSA) conjugates. R-SMCC-B (a conjugate of four RNase A attached with one BSA) and R-PEG-B (a mono-conjugate) were prepared using Sulfo-SMCC (a short bifunctional linker) and mal-PEG-NHS (a bifunctional PEG), respectively. Mal-PEG-NHS and hexadecylamine (HDA) were used to prepare the mono-conjugate, R-HDA-B, where HDA was adopted to bind BSA. The PEG linker can elongate the proximity between RNase A and BSA. In contrast, four RNase A were closely located on BSA in R-SMCC-B. R-SMCC-B showed the lowest Km and the highest relative enzymatic activity and kcat/Km in the three conjugates. Presumably, the tetravalent interaction of RNase A in R-SMCC-B can increase the binding affinity to its substrate. In addition, the slow release of BSA from R-HDA-B may increase the enzymatic activity of R-HDA-B. Our study is expected to provide strategies to develop protein–albumin conjugate with high therapeutic potential.
    No preview · Article · Dec 2012 · Journal of Biotechnology