Recent progress in recombinant DNA technology has allowed us to introduce genes for foreign useful proteins into microorganisms such as E.coli and to obtain their gene products in large amounts. In the present project, we aimed at, by controlling the properties of the expressed recombinant proteins to form inclusion bodies, increasing the expression efficiency of the recombinant proteins, suppressing their possible cell toxicity, and making their purification more feasible.
We used yeast mitochondrial MIF4p as a model recombinant protein with low expression efficiency in E.coli cells. We replaced the presequence of MIF4p with various peptides including SynA1, SynA2, SynB1, SynB2, SynC (derivatives of the preseuqnece of yeast cytochrome oxidase subunit IV) and T7TAG,and made corresponding fusion genes. We then introduced the fusion genes into E.coli cells and induced their overexpression. The expression levels of the fusion proteins were assessed by *mmunoblotting and compared with that of the mature, presequence-less MIF4p. MIF4p tagged with T7TAG and with SynB2 showed higher expression levels than the mature MIF4p. The Syn B2-MIF4p fusion protein was found in an inclusion body while the T7TAG-MIF4p fusion protein was recovered in a soluble fraction. These peptides (SynB2 and T7TAG) will offer a useful starting point to further optimize their abilities to increase expression levels of the attached proteins in E.coli cells.