THE MERCURY RESISTANCE OPERON OF THE INCJ PLASMID PMERPH EXHIBITS STRUCTURAL AND REGULATORY DIVERGENCE FROM OTHER GRAM-NEGATIVE MER OPERONS

The bacterial mercury resistance determinant carried on the Ind plasmi d pMERPH has been characterized further by DNA sequence analysis. From the sequence of a 4097 bp Bg/II fragment which confers mercury resist ance, it is predicted that the determinant consists of the genes merT, merP, merC and merA. The level of DNA sequence similarity between the se genes and those of the mer determinant of Tn21 was between 56.4 and 62.4%. A neighbour-joining phylogenetic tree of merA gene sequences w as constructed which suggested that pMERPH bears the most divergent Cr am-negative mer determinant characterized to date. Although the determ inant from pMERPH has been shown to be inducible, no regulatory genes have been found within the Bg/II fragment and it is suggested that a r egulatory gene may be located elsewhere on the plasmid. The cloned det erminant has been shown to express mercury resistance constitutively. Analysis of the pMERPH mer operator/promoter (O/P) region in vivo has shown constitutive expression from the mer P-TCPA promoter, which coul d be partially repressed by the presence of a trans-acting MerR protei n from a Tn21-like mer determinant. This incomplete repression of mer P-TCPA promoter activity may be due to the presence of an extra base b etween the -35 and -10 sequences of the promoter and/or to variation i n the MerR binding sites in the O/P region. Expression from the partia lly repressed mer P-TCPA promoter could be restored by the addition of inducing levels of Hg2+ ions. Using the polymerase chain reaction wit h primers designed to amplify regions in the merP and merA genes, 1.37 kb pMERPH-like sequences have been amplified from the Ind plasmid R39 1, the environmental isolate SE2 and from DNA isolated directly from n on-cultivated bacteria in River Mersey sediment. This suggests that pM ERPH-like sequences, although rare, are nevertheless persistent in nat ural environments.