Change of the Reactivity of Sulfhydryl Groups of the Membrane Proteins in Response to Amino Acids in Chang Liver Cells

Abstract

Intensity of fluorescence induced by reaction of a SH-directed fluorogenic reagent with whole Chang liver cells or their plasma membrane preparations which were previously treated with 0.5mM N-ethylmaleimide (NEM) plus 5mM leucine was enhanced by 30% or more in the presence of 5mM leucine or valine in assay medium consisting of Tris-Hepes buffer, pH 6.85, compared with the control. The dissociation constant for binding of leucine to the membrane protein (s) was estimated to be approximately 3mM from the increments of fluorescence intensity with increasing leucine concentration. On the other hand, the amount of [¹⁴C]NEM bound to the membrane preparations pretreated with NEM plus leucine was significantly reduced in the presence of leucine in the reaction medium. A decrease of reactivity of SH groups near the binding sites of the membrane proteins and of mobility of the fluorescent adducts on binding leucine was suggested to explain all these results consistently.