PMID:8995374
| Citation |
Liu, CE and Ames, GF (1997) Characterization of transport through the periplasmic histidine permease using proteoliposomes reconstituted by dialysis. J. Biol. Chem. 272:859-66 |
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| Abstract |
The superfamily of traffic ATPases (ABC transporters) includes bacterial periplasmic transport systems (permeases) and various eukaryotic transporters. The histidine permease of Salmonella typhimurium and Escherichia coli is composed of a membrane-bound complex containing four subunits and of a soluble receptor, the substrate-binding protein (HisJ), and is energized by ATP. The permease was previously reconstituted into proteoliposomes by a detergent dilution method (1). Here we extensively characterize the properties of this permease after reconstitution into proteoliposomes by dialysis and encapsulation of ATP or other reagents by freeze-thawing. We show that histidine transport depends entirely on both ATP and liganded HisJ, with apparent Km values of 8 mM and 8 microM, respectively, and is affected by pH, temperature, and salt concentration. Transport is irreversible and accumulation reaches a plateau at which point transport ceases. The permease is inhibited by ADP and by high concentrations of internal histidine. The inhibition by histidine implies that the membrane-bound complex HisQ/M/P carries a substrate-binding site. The reconstituted permease activity corresponds to about 40-70% turnover rate of the in vivo rate of transport. |
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| Keywords |
ATP-Binding Cassette Transporters; Adenosine Triphosphatases/metabolism; Amino Acid Transport Systems, Basic; Animals; Bacterial Proteins; Biological Transport, Active; Chromatography, Gel; Erythrocyte Membrane/metabolism; Glucosides/metabolism; Hydrogen-Ion Concentration; Liposomes/metabolism; Magnesium/metabolism; Membrane Proteins/metabolism; Membrane Transport Proteins/metabolism; Proteolipids/metabolism |
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