Expression of the gene encoding cytochrome c3 from the sulfate-reducing bacterium Desulfovibrio vulgaris in the purple photosynthetic bacterium Rhodobacter sphaeroides

Abstract

The gene encoding cytochrome c 3 ( cyc-gene) from Desulfovibrio vulgaris (Hildenborough) was cloned by G. Voordouw and S. Brenner (1986, Eur. J. Biochem. 159, 347–351) . The gene was expressed in Escherichia coli but only the apoprotein was observed (W. Pollock, P. Chemerika, M. Forrest, J. Beatty, and G. Voordouw, 1989, J. Gen. Microbiol. 135, 2319–2328) . In this study, the cyc-gene was cloned into the broad host range vector pRK404 and then introduced into the purple photosynthetic bacterium Rhodobacter sphaeroides. Cells grown anaerobically produced a significant amount of recombinant cytochrome c 3. The purified protein contains four hemes and the N-terminal protein sequence is identical to the published sequence of the native cytochrome c 3. Thus, R. sphaeroides was able to produce the mature cytochrome c 3 by combining the four steps of protein synthesis, exporting the protein across the membrane, cleaving the signal peptide, and inserting four hemes. It appears that the D. vulgaris promoter is not very efficiently used by R. sphaeroides. However, replacement of the promoter with a R. sphaeroides promoter should result in cytochrome c 3 overproduction.