The expression in Escherichia coli of a synthetic gene coding for the precursor of papain is prevented by its own putative signal sequence.

A 1048-bp gene coding for prepropapain was assembled from chemically synthesized oligodeoxyribonucleotides and cloned into a variety of Escherichia coli expression plasmids. We observed loss of plasmid when the preproP gene was expressed in E. coli either as the native precursor or fused at the C terminus of the first 592 amino acids (aa) of beta-galactosidase (beta Gal). Deletion of the putative 26-aa signal peptide (pre-region) increased plasmid stability. The level of maintenance for the different plasmid constructs correlated with the level of expression detected by immunoblotting. Constitutive expression of the beta Gal-propapain fusion generated insoluble granules in a protease-deficient E. coli host. The fusion protein was easily purified to near homogeneity by differential solubilization of the granules.