Cloning and Expression of Soluble Cytochrome c and Its Role in Polyvinyl Alcohol Degradation by Polyvinyl Alcohol-Utilizing Sphingopyxis sp. Strain 113P3(ENVIRONMENTAL BIOTECHNOLOGY)

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The gene encoding cytocrome c in the pva operon of Sphingopyxis sp. strain 113P3 was cloned, on the basis of the sequence of the gene for cytochrome c(GenBank accession no. AB190288). The deduced amino acid sequence of the gene showed homologies(37% and 47% identities) with two cytochromes c of different origins. The recombinant cytochrome c tagged with hexahistidines was expressed in the periplasm of Escherichia coll BL21(DE3) harboring pT-GroE, which was in accordance with the localization of cytochrome c in strain 113P3; the protein was purified to homogeneity. The purified recombinant cytochrome c was a monomeric protein with a molecular weight of 16.5 kDa. The oxidized and reduced forms of the protein showed absorption maxima at 409 nm and at 414, 520 and 550 nm, respectively. The recombinant cytochrome c was fully reduced by polyvinyl alcohol(PVA), coupled with a catalytic amount(1/10 molar concentration) of the recombinant PVA dehydrogenase(PVADH) of the same origin, suggesting that the cytochrome c involved in the pva operon is a physiological primary electron acceptor for PVADH and that PVA dehydrogenation is linked with the respiratory chain in Sphingopyxis sp. strain 113P3.
2008-02-25