Evidence for a second binding/transport site for chloride in erythrocyte anion transporter AE1 modified at glutamate 681.
Transport kinetics have been examined in erythrocyte anion transporter AE1 that has been chemically modified to convert glutamate 681 to an alcohol (E681OH AE1). Outward conductive Cl(-) flux in E681OH AE1 is inhibited by removal of extracellular Cl(-); this effect is the opposite of that in native AE1 and is consistent with coupled electrogenic 2:1 Cl(-)/Cl(-) exchange. A second Cl(-) binding/transport site is also suggested by the characteristics of (35)SO(4)(2-) flux in E681OH AE1: bilateral and cis Cl(-), which are normally inhibitory, accelerate (35)SO(4)(2-) flux. These effects would be expected if Cl(-) binds to a second transport site on SO(4)(2-)-loaded E681OH AE1, thereby allowing Cl(-)/SO(4)(2-) cotransport. Alternatively, the data can be explained without proposing Cl(-)/SO(4)(2-) cotransport if the rate-limiting event for (35)SO(4)(2-)/SO(4)(2-) exchange is external SO(4)(2-) release, and the binding of external Cl(-) accelerates SO(4)(2-) release. With either interpretation, these data indicate that E681OH AE1 has a binding/transport site for Cl(-) that is distinct from the main transport site. The effects of graded modification of E681 or inhibition by H(2)DIDS are consistent with the idea that the new Cl(-) binding site is on the same E681OH-modified subunit of the AE1 dimer as the normal transport site.
Jennings
Michael L
ML
Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA. JenningsMichaelL@uams.edu
D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry* / Q000378:metabolism; D000838:Anions; D001665:Binding Sites; D001692:Biological Transport; D001703:Biophysics / Q000379:methods*; D002244:Carbon / Q000737:chemistry; D002384:Catalysis; D002412:Cations; D002712:Chlorides / Q000737:chemistry*; D002713:Chlorine / Q000737:chemistry*; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004912:Erythrocytes / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents / Q000494:pharmacology; D007555:Isoxazoles / Q000494:pharmacology; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000737:chemistry; D008954:Models, Biological; D011485:Protein Binding; D011522:Protons; D013997:Time Factors
D001457:Anion Exchange Protein 1, Erythrocyte; D000838:Anions; D002412:Cations; D002712:Chlorides; D007202:Indicators and Reagents; D007555:Isoxazoles; D026901:Membrane Transport Proteins; D011522:Protons; C009686:N-ethyl-5-phenylisoxazolium-3'-sulfonate; D002713:Chlorine; D002244:Carbon