title: Erythrocyte Lii-Nao countertransport system. Inhibition by N-ethylmaleimide probes for a conformational change of the transport system.
abstract: Human erythrocytes were treated by a series of SH-reagents, including maleimides, iodo compounds, mercurials and oxidizing agents. Rates of Li efflux into Na-rich medium, Li leak and Lii-Nao countertransport were then determined. Of the 13 different reagents studied, only N-ethylmaleimide, iodoacetamide and iodoacetate inhibited selectively the countertransport activity. The effect of the various reagents indicates that the sensitive SH-groups of the countertransport system are not externally exposed. N-Ethylmaleimide was used to probe for changes elicited by substrate cations in Lii-Nao countertransport. In Na- and Li-free medium, inhibition of Lii-Nao countertransport by N-ethylmaleimide of 35% was reached within 2 s. In Na or Li medium, maximal inhibition was twice as great, but was attained much more slowly, within 10 min. Kinetic data and Hill plot analysis indicate the involvement of two classes of SH-groups: one expressed in the various media with and without substrate cations, and an additional one, which becomes specifically available to N-ethylmaleimide in the presence of external Na or Li. The affinity of Na to the site promoting inhibition by N-ethylmaleimide (apparent Km = 12 mM) is higher than the affinity of Na to its external countertransport site (apparent Km = 25 mM, as reported by Sarakadi, B., Alifimoff, J.K., Gunn, R.B. and Tosteson, D.C. (1978) J. Gen. Physiol. 72, 249-265). Reactivity of N-ethyl[14C]maleimide was not modified by the media tested. It is concluded that external Na and Li cause a conformational change in the protein(s) of the countertransport system in human erythrocytes.
mesh_terms: D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000097:blood*; D002412:Cations; D004912:Erythrocytes / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology*; D006801:Humans; D007460:Iodoacetamide / Q000494:pharmacology; D007461:Iodoacetates / Q000494:pharmacology; D019807:Iodoacetic Acid; D007700:Kinetics; D008094:Lithium / Q000097:blood* / Q000494:pharmacology; D011487:Protein Conformation; D012964:Sodium / Q000097:blood* / Q000494:pharmacology
chemical_list: D002352:Carrier Proteins; D002412:Cations; D007461:Iodoacetates; D008094:Lithium; D012964:Sodium; D005033:Ethylmaleimide; D019807:Iodoacetic Acid; D007460:Iodoacetamide