Cloning and sequencing of the gene encoding cytochrome c3 from Desulfovibrio vulgaris (Hildenborough)

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Abstract
The gene encoding the redox protein cytochrome c3 from Desulfovibrio vulgaris (Hildenborough) has been cloned using two synthetic oligonucleotides (one 17-mer and one 18-mer), designed to recognize the structural gene. Plasmid pCYC3 was derived from the clone and contains a 7.5 × 103-base EcoRI-HindIII insert of D. vulgaris DNA in pUC9. A 674-base-pair fragment of this insert was sequenced with the dideoxy-chain-termination procedure and found to contain the entire structural gene encoding cytochrome c3 bracketed by apparent Escherichia coli consensus sequences for initiation and termination of transcription. The amino acid sequence of 107 residues, derived from protein sequencing [Trousil, E. B. and Campbell, L. L. (1974) J. Biol. Chem. 249, 386–393], is confirmed by the nucleic acid sequence, which shows in addition that it is preceded by a hydrophobic, positively charged signal sequence of 21 residues. This amino-terminal extension functions in the export of cytochrome c3, which is thought to reside in the periplasm of D. vulgaris.