GROWTH-KINETICS OF ESCHERICHIA-COLI AND EXPRESSION OF A RECOMBINANT PROTEIN AND ITS ISOFORMS UNDER HEAT-SHOCK CONDITIONS

Preinduction culture conditions were found to have significant impact on the expression and post-translational modification of a recombinant human protein in Escherichia coli under heat shock conditions (30 to 42 degrees C shift). Higher preinduction growth rates (mu(g)) favored better cell viability, greater cell mass yields, and increased cloned gene expression during induction. Formation of recombinant protein iso forms (those containing N-epsilon-modified lysine residues) exhibited an increasing trend with increasing mu(g). The different extents of po st-translational modifications were suspected to be linked to the diff erent concentrations of certain heat shock protein chaperones resultin g from different mu(g). In view of the extensive involvement of E. col i heat shock proteins in cellular activities-including the synthesis, processing, modification, and degradation of proteins-at elevated temp eratures, it is believed that mu(g) dictated the cellular resources av ailable for synthesizing the heat shock proteins required for cell sur vival, which in turn determined the ability of the cells to respond to the heat shock. With a higher mu(g), both the synthesis of host prote ins (as indicated by cell growth and survival) and the cloned gene exp ression were enhanced. The results demonstrate that there exists an in termediate mu(g) for optimum production of the unmodified foreign prot ein in a heat shock environment. More importantly, they also illustrat e the feasibility of improving the recombinant protein homogeneity in fermentation, thereby facilitating downstream processing.