Pilot-scale production of human serum butyrylcholinesterase suitable for use as a bioscavenger against nerve agent toxicity
Pilot-scale production of human serum butyrylcholinesterase suitable for use as a bioscavenger against nerve agent toxicity
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DOI：
10.1016/j.procbio.2010.04.021
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发表时间：
2010-08-01
影响因子：
4.4
通讯作者：
Doctor, Bhupendra P.
中科院分区：
文献类型：
--
作者：
Saxena, Ashima;Tipparaju, Prasanthi;Doctor, Bhupendra P.
人血清丁酰胆碱酯酶(Hu BChE)是目前预防有机磷(OP)神经毒剂毒性最合适的候选药物。据估计，200毫克的剂量将保护人类免受2×LD(50)的梭曼的伤害，这意味着人体临床研究需要克量的酶。为此，我们报道了适合于从Cohn组分IV-4糊状物中中试提纯Hu BChE的第一步工艺的开发。这一过程包括Cohn组分IV-4糊剂的再悬浮，然后是普鲁卡因胺亲和和DEAE阴离子交换层析。该方法从80公斤的Cohn组分IV-4糊状物中获得6-7g(430万-500万U)的纯化酶。该酶的比活力接近700U/mg，SDS-PAGE显示一条亚基相对分子质量为85 kDa的主带，纯度为97%。该工艺收率高、纯度高，适合于临床前期生产。爱思唯尔有限公司出版。
Human serum butyrylcholinesterase (Hu BChE) is currently the most appropriate candidate for the prophylaxis of humans against organophosphate (OP) nerve agent toxicity. It is estimated that a dose of 200 mg will protect a human against 2 x LD(50) of soman, which means that gram quantities of enzyme are needed for human clinical studies. Toward this effort, we report the development of the first procedure that is suitable for the pilot-scale purification of Hu BChE from Cohn fraction IV-4 paste. This procedure involved resuspension of Cohn fraction IV-4 paste, followed by procainamide affinity and DEAE anion-exchange chromatography. The procedure yielded 6-7 g (4.3-5 million U) of purified enzyme from 80 kg of Cohn fraction IV-4 paste. The enzyme was >97% pure as judged by a specific activity of similar to 700 U/mg and a major band with a subunit molecular weight of 85 kDa on SDS-PAGE. The high yield and purity obtained suggest that this manufacturing procedure is suitable for the pre-clinical production of Hu BChE. Published by Elsevier Ltd.