The Multidrug Transporter LmrP Protein Mediates Selective Calcium Efflux

Multidrug Transport Protein NorM from Vibrio cholerae Simultaneously Couples to Sodium- and Proton-Motive Force

“…In recent studies on the proton/drug antiporter LmrP from L. lactis, for example, Asp-142 in the N-terminal half was found to act as a dedicated proton-binding site that facilitates conformational changes underlying the transport cycle. In addition, Asp-235 and Glu-327 in the C-terminal half were found to interact with protons and to facilitate the binding of cationic substrates (12,25,27). On the other hand, in the crystallized bacterial sodium-leucine symporter LeuT, one binding site for dehydrated Na ϩ is formed by a carboxylate moiety and neighboring main chain carbonyl and side chain amide atoms (33).…”

“…For this purpose, membrane vesicles (10 g of total membrane protein, see below) were mixed with 30 l of 3ϫ sample loading buffer containing 6% (w/v) SDS, then mixed with DTT (1 mM final concentration), and subsequently incubated at 20°C for 10 min before loading on gel (5 g of protein/lane). Inside-out membrane vesicles were prepared from washed E. coli cells (26) in 100 mM potassium P i buffer (pH 7.0) using a Basic Z cell disruptor (Constant Systems) (27). Membrane vesicles were solubilized for 4 h in the presence of 1% (w/v) DDM in 50 mM potassium P i (pH 8.0), 200 mM K 2 SO 4 , 5% glycerol (v/v), 30 mM imidazole and EDTA-free protease inhibitor (Roche Applied Science).…”

Molecular Disruption of the Power Stroke in the ATP-binding Cassette Transport Protein MsbA

“…However, all three alanine mutants displayed greatly reduced ATP-dependent Hoechst 33342 transport activities in ISOVs compared with the WT protein, in an assay in which the fluorescence quenching of the dye was detected following its transport from the membrane into the acidic, aqueous lumen of the inside-out membrane vesicles (Fig. 3A) (32,34). Consistent with this observation, we also found that each alanine mutant was defective in mediating metabolic energy-dependent export of the cationic substrate ethidium from cells, in an assay in which the fluorescence quenching of the dye was detected following its dissociation from genomic DNA during transport (Fig.…”

Functional Linkage between Genes That Regulate Osmotic Stress Responses and Multidrug Resistance Transporters: Challenges and Opportunities for Antibiotic Discovery

“…1B). Examples of MDR transporters with alternative functions include the MFS proton/ drug antiporter MdfA and MdtM from E. coli, which have been found to function in alkaline pH homeostasis by exchanging Naϩ or Kϩ for Hϩ (26,27); PsmrAB, a SMR protein from halophilic bacteria that functions as a novel two-component Naϩ/Hϩ antiporter (28); and LmrP, an MFS proton/cationic drug antiporter from Lactococcus lactis that can also mediate calcium efflux (29).…”