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Cell-free production of functional antibody fragments

机译:无细胞产生功能性抗体片段

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Botulinum neurotoxin serotype B (BoNT/B)-specific Fab was expressed in a cell-free protein synthesis system derived from an E. coli extract. The cell-free synthesized antibody fragment was found to be effective in neutralizing the toxicity of BoNT/B in animal studies. Expression of functional Fab required an appropriately controlled and stably maintained redox potential. Under an optimized redox condition, the cell extract, whose disulfide reducing activity had been exhausted, could generate bio-functional Fab molecules. Use of a cell extract enriched with molecular chaperones (GroEL/ES) and disulfide bond isomerases were effective in obtaining larger quantities of functional Fab. Under the optimized reaction conditions, approximately 30 μg of functional Fab was obtained after purification from 1 mL reaction mixture.
机译:肉毒杆菌神经毒素血清型B(BoNT / B)特异性Fab在源自大肠杆菌提取物的无细胞蛋白质合成系统中表达。在动物研究中,发现无细胞的合成抗体片段可有效中和BoNT / B的毒性。功能性Fab的表达需要适当控制和稳定维持的氧化还原电位。在优化的氧化还原条件下,已耗尽其二硫键还原活性的细胞提取物可以产生具有生物功能的Fab分子。使用富含分子伴侣(GroEL / ES)和二硫键异构酶的细胞提取物可有效获得大量功能性Fab。在优化的反应条件下,从1 mL反应混合物中纯化后,可获得约30μg的功能性Fab。

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