MUTANTS OF THE BACILLUS-SUBTILIS MULTIDRUG TRANSPORTER BMR WITH ALTERED SENSITIVITY TO THE ANTIHYPERTENSIVE ALKALOID RESERPINE

The Bacillus subtilis multidrug transporter Bmr effluxes structurally diverse toxic compounds out of bacterial cells. Antihypertensive alkal oid reserpine reverses Bmr-mediated multidrug resistance by inhibiting drug transport. We have obtained a mutant of the bmr gene that provid es a normal level of multidrug resistance, which can, however, only be reversed by very high concentrations of reserpine. Reduction of Bmr s ensitivity to reserpine has been caused by the substitution of Leu for Val286 in the Bmr molecule. This mutation also led to a dramatic decr ease of [H-3]reserpine binding to membrane vesicles prepared from the Bmr-overexpressing bacteria. Leucine is larger than valine by one meth ylene group. Substitution of Val286 with a smaller residue, glycine, h ad an opposite effect. It led to increased sensitivity of Bmr to reser pine and increased affinity of reserpine binding to the membranes prep ared from Bmr-overexpressing bacteria. Neither of the mutations signif icantly changed the sensitivity of Bmr to rescinnamine, a structural a nalog of reserpine. The results suggest that Val286 is involved in the formation of the reserpine-binding site of the Bmr molecule.