San K Y, Bennett G N, Aristidou A A, Chou C H
Department of Chemical Engineering, Rice University Houston, TX 77251-1892.
Ann N Y Acad Sci. 1994 May 2;721:257-67. doi: 10.1111/j.1749-6632.1994.tb47399.x.
The accumulation of acetate is one of the most commonly encountered problems in attaining high levels of recombinant protein production using E. coli. Two different approaches are examined to reduce the rate of acetate formation. The effects of reduced acetate accumulation on recombinant protein production were also investigated. In the first approach, E. coli mutant strains deficient in enzymes involved in the acetate synthesis pathways were isolated and characterized. The level of specific production of beta-galactosidase by the mutant strain is three times higher than its parent strain. In another approach, metabolic engineering techniques were employed to fine-tune the central metabolic pathways to reduce the amount of acetate formation. The resulting strain, which carries the acetolactase synthase gene from B. subtilis, is successful in maintaining a very low level of acetate accumulation. The ALS-containing strain is also capable of producing higher levels of recombinant protein than its parent strain.
在利用大肠杆菌实现高水平重组蛋白生产的过程中，乙酸盐的积累是最常见的问题之一。研究了两种不同的方法来降低乙酸盐的形成速率。还研究了减少乙酸盐积累对重组蛋白生产的影响。在第一种方法中，分离并鉴定了缺乏参与乙酸盐合成途径的酶的大肠杆菌突变菌株。突变菌株β-半乳糖苷酶的比生产水平比其亲本菌株高3倍。在另一种方法中，采用代谢工程技术对中心代谢途径进行微调，以减少乙酸盐的形成量。所得菌株携带来自枯草芽孢杆菌的乙酰乳酸合酶基因，成功地保持了极低水平的乙酸盐积累。含有ALS的菌株也能够比其亲本菌株产生更高水平的重组蛋白。
