The mercury resistance operon of the IncJ plasmid pMERPH exhibits structural and regulatory divergence from other Gramnegative mer operons

Abstract

Summary: The bacterial mercury resistance determinant carried on the IncJ plasmid pMERPH has been characterized further by DNA sequence analysis. From the sequence of a 4097 bp Bg/II fragment which confers mercury resistance, it is predicted that the determinant consists of the genes merT, merP, merC and merA. The level of DNA sequence similarity between these genes and those of the mer determinant of Tn21 was between 56.4 and 62.4%. A neighbourjoining phylogenetic tree of merA gene sequences was constructed which suggested that pMERPH bears the most divergent Gram-negative mer determinant characterized to date. Although the determinant 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 regulatory gene may be located elsewhere on the plasmid. The cloned determinant 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 could be partially repressed by the presence of a trans-acting MerR protein 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 between the −35 and −10 sequences of the promoter and/or to variation in the MerR binding sites in the O/P region. Expression from the partially repressed mer P_TCPA promoter could be restored by the addition of inducing levels of Hg²⁺ ions. Using the polymerase chain reaction with primers designed to amplify regions in the merP and merA genes, 1.37 kb pMERPH-like sequences have been amplified from the IncJ plasmid R391, the environmental isolate SE2 and from DNA isolated directly from non-cultivated bacteria in River Mersey sediment. This suggests that pMERPH-like sequences, although rare, are nevertheless persistent in natural environments.