MUTATIONS AFFECTING TRANSLATIONAL COUPLING BETWEEN THE REP GENES OF AN INCB MINIPLASMID

The nature of translational coupling between repB and repA, the overlapping rep genes of the IncB plasmid pMU720, was examined. Mutations in the start codon of the promoter proximal gene, repB, reduced the efficiency of translation of both rep genes. Moreover, there was no independent initiation of repA translation in the absence of repB translation. The position of the repB stop codon was crucial for the efficient expression of repA, with the wild-type positioning being optimal. Translational coupling was found to be totally dependent on the formation of a pseudoknot structure. A model which invokes formation of a pseudoknot to facilitate initiation of repA is proposed.