THE DESULFUROMONAS-ACETOXIDANS TRIHEME CYTOCHROME C(7) PRODUCED IN DESULFOVIBRIO-DESULFURICANS RETAINS ITS METAL REDUCTASE-ACTIVITY

Multiheme cytochrome c proteins that belong to class III have been rec ently shown to exhibit a metal seductase activity, which could be of g reat environmental interest, especially in metal bioremediation. To ge t a better understanding of these activities, the gene encoding cytoch rome c(7) from the sulfur-reducing bacterium Desulfuromonas acetoxidan s was cloned from genomic DNA by PCR and expressed in Desulfovibrio de sulfuricans G201. The expression system was based on the cyc transcrip tion unit from Desulfovibrio vulgaris Hildenborough and led to the syn thesis of holocytochrome c(7) when transferred by electrotransformatio n into the sulfate reducer Desulfovibrio desulfuricans G201. The produ ced cytochrome was indistinguishable from the proton purified from Des ulfuromonas acetoxidans cells with respect to several biochemical and biophysical criteria and exhibited the same metal reductase activities as determined from electrochemical experiments. This suggests that th e molecule was correctly folded in the host organism. Desulfovibrio de sulfuricans produces functional multiheme c-type cytochromes from bact eria belonging to a different genus and may be considered a suitable h ost for the heterologous biogenesis of multiheme c-type cytochrome for either structural or engineering studies. This report, which presents the first example of the transformation of a Desulfovibrio desulfuric ans strain by electrotransformation, describes work that is the first necessary step of a protein engineering program that aims to specify t he structural features that are responsible for the metal reductase ac tivities of multiheme cytochrome c(7).