7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-C12-sphingomyelin/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-C12-sphingomyelin/out - ? 414301 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-glucosylceramide/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-glucosylceramide/out - ? 414302 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-lactosylceramide/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-lactosylceramide/out - ? 414303 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylcholine (16:0, 6:0)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylcholine (16:0, 6:0)/out - ? 414304 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine (16:0, 6:0)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species. Translocation of (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine is inhibited by the presence of the putative physiological substrate lipid A Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine (16:0, 6:0)/out - ? 414305 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine (18:1)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species. Translocation of (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine is inhibited by the presence of the putative physiological substrate lipid A Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylethanolamine (18:1)/out - ? 414306 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylglycerol (16:0, 6:0)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylglycerol (16:0, 6:0)/out - ? 414307 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylserine (16:0, 6:0)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylserine (16:0, 6:0)/out - ? 414308 7.5.2.6 ATP + H2O + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylserine (18:1)/in bacterial membrane vesicles isolated from Escherichia coli overexpressing MsbA display ATP-dependent translocation of several fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labelled phospholipid species Escherichia coli ADP + phosphate + (7-nitrobenz-2-oxa-1,3-diazole)-phosphatidylserine (18:1)/out - ? 414309 7.5.2.6 ATP + H2O + daunorubicin[side 1] - Salmonella enterica subsp. enterica serovar Typhimurium ADP + phosphate + daunorubicin[side 2] - ? 444993 7.5.2.6 ATP + H2O + DMSO/in - Escherichia coli ADP + phosphate + DMSO/out - ? 433453 7.5.2.6 ATP + H2O + ethidium/in - Escherichia coli ADP + phosphate + ethidium/out - ? 433456 7.5.2.6 ATP + H2O + Hoechst 33342 - Escherichia coli ADP + phosphate + Hoechst 33342/out - ? 433465 7.5.2.6 ATP + H2O + Hoechst 33342/in - Escherichia coli ADP + phosphate + Hoechst 33342/out - ? 414320 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Salmonella enterica subsp. enterica serovar Typhimurium ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Escherichia coli ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Lactococcus lactis subsp. lactis ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa ATCC 15692 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Lactococcus lactis subsp. lactis NZ9000 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa 1C ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa PRS 101 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa DSM 22644 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa CIP 104116 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa LMG 12228 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Escherichia coli K12 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Escherichia coli W3110 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + lipid A-core oligosaccharide[side 1] - Pseudomonas aeruginosa JCM 14847 ADP + phosphate + lipid A-core oligosaccharide[side 2] - ? 446167 7.5.2.6 ATP + H2O + Lipid A/in - Escherichia coli ADP + phosphate + Lipid A/out - ? 376343 7.5.2.6 ATP + H2O + Lipid A/in substrate binding structure, overview Escherichia coli ADP + phosphate + Lipid A/out - ? 376343 7.5.2.6 ATP + H2O + Lipid A/in MsbA contains two substrate-binding sites that communicate with both the nucleotide-binding domain and with each other, one is a high affinity binding site for the physiological substrate, lipid A, and the other site interacts with drugs with comparable affinity, overview Salmonella enterica subsp. enterica serovar Typhimurium ADP + phosphate + Lipid A/out - ? 376343 7.5.2.6 ATP + H2O + lipo-oligosaccharide/in - Pseudomonas aeruginosa ADP + phosphate + lipo-oligosaccharide/out - ? 141472 7.5.2.6 ATP + H2O + lipo-oligosaccharide/in MsbA protein shown as a member of the ABC transporter family, differences to MsbA protein of Escherichia coli determined Pseudomonas aeruginosa ADP + phosphate + lipo-oligosaccharide/out - ? 141472 7.5.2.6 ATP + H2O + lipopolysaccharide/in - Salmonella enterica subsp. enterica serovar Typhimurium ADP + phosphate + lipopolysaccharide/out - ? 402473 7.5.2.6 ATP + H2O + lipopolysaccharide/in - Escherichia coli ADP + phosphate + lipopolysaccharide/out - ? 402473 7.5.2.6 ATP + H2O + lipopolysaccharide/in - Aspergillus nidulans ADP + phosphate + lipopolysaccharide/out - ? 402473 7.5.2.6 ATP + H2O + lipopolysaccharide/in MsbA is an essential ABC transporter involved in lipid A transport across the cytoplasmic membrane Salmonella enterica subsp. enterica serovar Typhimurium ADP + phosphate + lipopolysaccharide/out - ? 402473 7.5.2.6 ATP + H2O + lipopolysaccharide/in - Aspergillus nidulans TN02A3 ADP + phosphate + lipopolysaccharide/out - ? 402473 7.5.2.6 ATP + H2O + phosphoethanolamine[side 1] fluorescently NBD (7-nitrobenz-2-oxa-1,3-diazole)-labeled Escherichia coli ADP + phosphate + phosphoethanolamine[side 2] - ? 447098 7.5.2.6 ATP + H2O + verapamil/in - Escherichia coli ADP + phosphate + verapamil/out - ? 376359 7.5.2.6 additional information MsbA functions as an ATP-dependent lipid translocase that transports lipid A from the inner to the outer leaflet of the cytoplasmic membrane. MsbA is able to hydrolyze TNP-ATP, albeit at a lower rate than its corresponding ATPase activity Escherichia coli ? - ? 89 7.5.2.6 additional information MsbA is an essential ABC transporter in Gram-negative bacteria Escherichia coli ? - ? 89 7.5.2.6 additional information MsbA is an essential ABC transporter in Gram-negative bacteria Salmonella enterica subsp. enterica serovar Typhimurium ? - ? 89 7.5.2.6 additional information the MsbA2 protein in the inner membrane is essential to ensure the symbiotic interaction with the host plant alfalfa. Sinorhizobium meliloti invades plant cells via plant-derived structures known as infection threads. However, MsbA2 is not essential for the membrane transport of either lipopolysaccharide or phospholipids in the organism, but in the absence of MsbA2 the polysaccharide content of Sinorhizobium meliloti is altered Sinorhizobium meliloti ? - ? 89 7.5.2.6 additional information binding of of amphipathic drugs, e.g. of daunorubicin, quercetin, verapamil, or propafenone GP12, alters the protein conformation Salmonella enterica subsp. enterica serovar Typhimurium ? - ? 89 7.5.2.6 additional information concerted conformational rearrangements occur during MsbA ATPase cycle Escherichia coli ? - ? 89 7.5.2.6 additional information MsbA cannot efficiently transport a substrate lacking phosphorylation at the 4'-position of lipid A Salmonella enterica subsp. enterica serovar Typhimurium ? - ? 89 7.5.2.6 additional information MsbA cannot efficiently transport a substrate lacking phosphorylation at the 4'-position of lipid A Escherichia coli ? - ? 89 7.5.2.6 additional information purified MsbA from Escherichia coli displays high ATPase activity, and binds to lipids and lipid-like molecules, including lipid A, with affinity in the low micromolar range. Purified MsbA is reconstituted into proteoliposomes of Escherichia coli lipid and shows ability to translocate 7-nitrobenz-2-oxa-1,3-diazole (NBD)-labeled lipid derivatives. In this system, the protein displays maximal lipid flippase activity of 7.7 nmol of lipid translocated per mg of protein over a 20 min period for an acyl chain-labeled phosphatidylethanolamine derivative. Lipid flippase activity requires ATP hydrolysis, and is dependent on the concentration of ATP and NBD-lipid. MsbA can accommodate lipids with bulky carbohydrate headgroups Escherichia coli ? - ? 89 7.5.2.6 additional information simultaneous high affinity binding to MsbA of lipid A (the putative physiological substrate) and daunorubicin, which suggests that the protein has separate binding sites for these two compounds. The effects of nucleotide and lipid A/daunorubicin binding to MsbA are additive, and binding could occur in any order. The two substrate-binding sites appear to communicate with each other, and also with the nucleotide-binding site in the nucleotide-binding domains, NBDs. The two monomers function independently, and do not interact co-operatively with each other, analysis using MIANS-labeled enzyme, MIANS, i.e. 2-(4-maleimidylanilino)naphthalene-6-sulfonic acid, is a cysteine-reactive fluorescent probe for soluble and membrane-bound proteins, fluorescence quenching studies. The percentage quenching values observed for lipid A and daunorubicin are also similar regardless of the order of titration. The binding affinity for lipid A is reduced about 5fold at 23°C and about 7fold at 10°C when the daunorubicin-binding site is occupied first Salmonella enterica subsp. enterica serovar Typhimurium ? - ? 89