Involvement of sulfhydryl groups in the action of the insulin and radiation on thymocyte Na+-dependent amino acid transport.
1,p-Chloromercuribenzene sulfonate concentrations less than 10(-5) M stimulate the uptake by thymocytes of 2-aminoisobutyrate, a non-metabolized amino acid. At concentrations greater than 10(-5) M of this reagent, transport is impaired and cell viability is effected. In contrast, 5,5'-dithiobis-(2-nitrobenzoate) between 10(-4) and 10(-6) M produces only stimulation of 2-aminoisobutyrate uptake after treating for 10 min. 2. Treatment of thymocytes with 10(-4)M 5,5'-dithiobis-(2-nitrobenzoate) reveals at least three categories of reactive SH groups. Titration of the most rapidly reacting category, 4 - 10(7)-7 - 10(7)/cell, activates 2-aminoisobutyrate transport to the same extent as does p-chloromercuribenzene sulfonate. Cells treated with 10(-6) M insulin showed a 30-50% reduction in the number of sulfhydryl groups that could be titrated with 5,5'-dithiobis-(2-nitrobenzoate). In thymocytes treated with 10(-6) M p-chloro(203Hg)mercuribenzene sulfonate, addition of 10(-6) or 10(-9) M insulin before treatment with the sulhydryl reagent again reduces the number of titrable SH groups by 20%. 3. Insulin (10(-10)-10(-6) M) also stimulates 2-aminoisobutyrate uptake, but the effects of insulin and SH blocker are not additive. 4. Insulin, but not p-chloromercuribenzene sulfonate, prevents the impairment of 2-aminoisobutyrate transport caused by gamma-irradiation. Treatment of cells with p-chloromercuribenzene sulfonate prior to irradiation increases the radiation impairment of 2-aminoisobutyrate transport. 5. gamma-irradiation reduces the number of 5,5'-dithiobis-(2-nitrobenzoate) reactive sulfhydryl residues by 37%. 6. A model for the action of insulin and irradiation on 2-aminoisobutyrate transport is presented.
D002731:4-Chloromercuribenzenesulfonate / Q000494:pharmacology; D000621:Aminoisobutyric Acids / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D001692:Biological Transport; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D005260:Female; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D011485:Protein Binding; D011487:Protein Conformation; D011830:Radiation Effects; D051381:Rats; D012964:Sodium / Q000494:pharmacology*; D013438:Sulfhydryl Compounds / Q000032:analysis; D013950:Thymus Gland / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects
D000621:Aminoisobutyric Acids; D007328:Insulin; D013438:Sulfhydryl Compounds; D002731:4-Chloromercuribenzenesulfonate; D004228:Dithionitrobenzoic Acid; D012964:Sodium