表达rAPC大肠杆菌的高密度发酵及纯化产物的抑瘤活性

High Cell Density Fermentation of a Recombinant E.coli Strain P_1 Producing an Anti-S_ (180) Tumor Recombinant Protein(rAPC)

重组菌的高密度发酵技术是高效表达异源蛋白的一项重要技术。本文研究了培养基组成、培养条件以及诱导条件等对最终的菌体密度和rAPC表达量的影响 ,确立了最优的高密度发酵条件。此外 ,还探讨了不同补料流加方式对菌体生长和rAPC表达的影响 ,结果表明DO -stat补料流加方式具有显著增加菌体量和重组蛋白表达量的作用。发酵 16小时后 ,菌体密度可达OD60 0 10 6,每升发酵液中rAPC含量可达 3 5 2 g。用纯化好的rAPC对皮下接种S180 的小鼠灌胃或腹腔注射 ,剂量为每天 3 4mg/kg～ 13 4mg/kg ,共10天 ,结果表明rAPC对小鼠S180 肉瘤有显著的抑制作用 ,瘤重抑瘤率分别在 4 5 % 64%之间 ,且对荷瘤小鼠的白细胞数量和胸腺指数均无明显影响。

High cell density fermentation of a recombinant E.coli strain P1 was studied. Several factors including medium composition, cultivation conditions and induction conditions that influenced the final cell density and rAPC(recombinant allophycocyanin) production was optimized. The effects of three feeding strategies were also investigated. The result showed that DO-stat feeding strategy had significant effect in both enhancing final cell density and promoting rAPC expression. Under the optimized fermentation condition, a cell density of OD 600106 was reached after 16 hours of fermentation and the expressed rAPC reached to a content of about 3.52g per litre culture broth. S 180 tumor carrying mice were then treated with purifed rAPC ig or ip 3.4mg/kg～13.4mg/kg per day for ten days. The results showed that rAPC could decrease weight of S 180 tumor significantly. The inhibitory rate ranged from 45% to 64% and rAPC had no negative effect on thymus index and leukocyte count of the test mice.