San, Ka-Yiu2009-06-042009-06-041995Chou, Chih-Hsiung. "Biochemical and genetic engineering strategies to enhance recombinant protein production in Escherichia coli." (1995) Diss., Rice University. <a href="https://hdl.handle.net/1911/16806">https://hdl.handle.net/1911/16806</a>.https://hdl.handle.net/1911/16806Escherichia coli has been widely used to produce high-value recombinant proteins for years. Although high recombinant protein productivity can be attained, perhaps the most important goal for such processes, that of achieving both high gene expression and high cell density simultaneously, is still challenging to both biochemists and biochemical engineers. A series of approaches to overcome this problem are evaluated in this study. First, a novel pH-inducible gene expression system, in which the expression of foreign gene products is directed by a pH down-shift, was chosen and characterized. This system was shown to have many attractive features, including high-level expression (40% of total cellular protein), fast response, and easy manipulation. It thus can serve as a proper model system for studying the fundamental mechanism of high-level gene expression in E. coli. Second, several factors limiting the culture performance were identified by systematically optimizing the culture conditions. Among those, acetate overproduction was shown to be critically involved. Various approaches on the basis of biochemical and genetic engineering techniques were successfully exploited to bypass such a cultivation bottleneck. Finally, the effects of various genetic elements, including the genes responsible for carbohydrate uptake and several stationary-phase genes, on recombinant protein production were investigated by genetic manipulation of the host strain. Several strategies were then developed to genetically construct more potent strains for recombinant protein production. The information is important not only for the modification of several structural models developed recently, but also for economic interest in terms of improving bioprocesses without further investment in equipment.98 p.application/pdfengCopyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.Chemical engineeringThesisTHESIS CH.E. 1995 CHOU