title: Evidence for a second binding/transport site for chloride in erythrocyte anion transporter AE1 modified at glutamate 681.
abstract: 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.
authors.affiliation: Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA. JenningsMichaelL@uams.edu
authors.forename: Michael L
authors.initials: ML
authors.lastname: Jennings
mesh_terms: 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
chemical_list: 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