ATP does not regulate the reconstituted glucose transporter.
ATP has been reported to affect glucose transport in human erythrocytes and resealed erythrocyte ghosts [Jacquez, J. A. (1983) Biochim. Biophys. Acta 727, 367-378; Jensen, M. R., & Brahm, J. (1987) Biochim. Biophys. Acta 900, 282-290]. In more detailed studies, effects of micromolar levels of ATP on transport in ghosts and inside-out vesicles, and on the fluorescence of ghosts and the purified glucose transporter [Carruthers, A. (1986) Biochemistry 25, 3592-3602; Hebert, D. N., & Carruthers, A. (1986) J. Biol. Chem. 261, 10093-10099; Carruthers, A. (1986) J. Biol. Chem. 261, 11028-11037], have been interpreted as supporting a model in which ATP regulates the catalytic properties of the transporter. Both allosteric and covalent effects of ATP were proposed; among the allosteric effects was a 60% reduction in the Km for zero-trans uptake. In order to test whether allosteric ATP regulation of the transporter occurs, we reconstituted glucose transport activity into liposomes using erythrocyte membranes without detergent treatment. The effects of ATP, present either outside, inside, or both inside and outside the liposomes, on the transport activity were examined. Effects of ATP on trypsin-treated liposomes, which have only a single orientation of active transporters, were also tested. While the model predicts activation by ATP, only inhibition was observed. This was significant only at millimolar concentrations of ATP, in contrast to the previously reported effects at micromolar levels, and was primarily on the extracellular surface of the transporter. In addition, the ATP effects on reconstituted transport were nonspecific, with similar effects produced by tripolyphosphate.(ABSTRACT TRUNCATED AT 250 WORDS)
Wheeler
T J
TJ
Department of Biochemistry, University of Louisville School of Medicine, Kentucky 40292.
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D001693:Biological Transport, Active / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008081:Liposomes; D008274:Magnesium / Q000494:pharmacology; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D011122:Polyphosphates / Q000494:pharmacology
D008081:Liposomes; D009004:Monosaccharide Transport Proteins; D011122:Polyphosphates; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D008274:Magnesium; D005947:Glucose; C005692:triphosphoric acid; D002118:Calcium