Enhancement of extracellular production of a Cellulomonas fimi exoglucanase in Escherichia coli by the reduction of promoter strength

Extracellular Expression, Purification, and Characterization of a Winter Flounder Antifreeze Polypeptide from Escherichia coli

“…Plasmid tacIQpar8cex construct, a excretion vector used to express/excrete a Cellulomonas fimi exoglucanase (17), was selected to be the parental plasmid for the construction of the rAFP expression/excretion plasmid, placIQpar8AF. Enzymes and chemicals used in DNA manipulation were purchased from Pharmacia, Sigma, and Bio-Rad.…”

“…However, to overcome the small molecular size of winter flounder AFPs, the AFPs were produced as chimeric proteins, AFP multimers, or AFP with multiple 11-amino acid repeats (10 -14). Recent achievements in the engineering of E. coli heterologous protein excretion systems (15)(16)(17) have provided an attractive and alternative approach to the construction of recombinant strains for the large-scale production of secreted AFPs, which can be easily purified from culture medium. Moreover, such a strain may facilitate mutagenic manipulation to produce AFP mutants.…”

Heterologous Expression and Extracellular Secretion of Cellulases in Recombinant Microbes

“…The cellobiose metabolic operon from Klebsiella oxytoca has been introduced into E. coli, but the expression level of the cellobiose transporter and metabolic genes was poor, and hence could not support the growth of E. coli on cellobiose (Moniruzzaman et al 1997). Cellulases from several species of Clostridium, Bacillus, Cellulomonas, and Ruminococcus have been expressed and characterized in E. coli (Hinchliffe 1984;Zappe et al 1986;Fierobe et al 1991;ReverbelLeroy et al 1996;Lam et al 1997;ReverbelLeroy et al 1997;Lee et al 2008;Li et al 2009). Co-expression of endoglucanase from B. pumilus and β-glucosidase from Fervidobacterium spp.…”

Applications, and Efficient Large-Scale Production, of Recombinant Human Epidermal Growth Factor

“…The vector developed (Lam etat., 1997) is a high copy-number plasmid containing (a) a cassette into which foreign DNA may be inserted, and (b) the lacl Q gene. The cassette (Figure 3.1A) contains 1J first, the lacUV5 promoter, which was deliberately chosen to be weaker than the tac promoter used originally (Wong and Sutherland, 1993; Wong et at.~1998a) as (over)...expression of cloned genes from the tac promoter sometimes results in death of recombinant cells (Lam et al, 1997). The lac operator follows.…”

“…The enzymatic degradation of waste cellulosic products, to make fuel alcohol t is a desirable biotechnological goal, and recombinant E. coli strains have been constructed which excrete all 3 of the required cellulosic enzymes (Lam et al, 1997;Wong et a/., 1998a). High-level production of fish antifreeze peptides is required by the dairy industry (the peptides will be incorporated into frozen products to improve 62 W,,-K.R.…”