Effect of Immobilization on Metabolism of Escherichia Coli: Research Planning Grant

在过去的几年里，使用固定化的全细胞进行生化转化的情况显著增加。最重要的领域之一是固定化对所用细胞内在新陈代谢的影响。关于固定化如何影响碳流、能量产生和产物形成的报道很少，特别是在大肠杆菌方面。初步结果表明，固定化后，新陈代谢发生了显著变化。在藻酸盐中生长的大肠杆菌细胞每个细胞合成明显更多的β-半乳糖苷酶，与游离细胞相比，正常代谢副产物醋酸盐、丙酮酸和乳酸的数量增加。这些结果支持这样一种假设，即固定改变了碳消耗和增长的相对速度。这项工作的具体目标是检验以下两个假设：i)固定化细胞消耗的多余碳被转移与酸性副产物或诱导蛋白的合成有关；以及ii)酸副产物去除过程中的传质限制导致了观察到的代谢变化。最终目标是利用固定化诱导的代谢变化来过量生产重组蛋白。

The use of immobilized whole cells to conduct biochemical conversions has increased significantly in the last several years. One of the most important areas is the effects of immobilization on the intrinsic metabolism of the cells employed. Little has been reported, particularly with respect to E. coli, on how immobilization affects carbon flow, energy production, and product formation. Preliminary results have shown that significant changes in metabolism occur upon immobilization. E. coli cells grown in alginate synthesize significantly more beta-galactosidase per cell, and elevated quantities of the normal metabolic byproducts acetate, pyruvate, and lactate, compared to free cells. These results support the hypothesis that immobilization changes the relative rates of carbon consumption and growth. The specific objectives of this work are to test the following two hypotheses: i) that the excess carbon consumed by immobilized cells is diverted to the synthesis of acid byproducts or of inducible protein; and ii) that mass transfer limitations on the removal of acid byproducts cause the observed changes in metabolism. The ultimate goal is to exploit immobilization-induced metabolic changes for the overproduction of recombinant proteins.