Differential acid stabilities of citraconylated amino groups of glucagon. Preparation of N alpha-citraconyl glucagon and evaluation of its biological properties.
Acylation of the alpha- and epsilon-amino groups of histidine-1 and lysine-12 in glucagon with citraconic anhydride resulted in the formation of amide bonds which displayed different stabilities to hydrolysis under mild acid conditions. Treatment of N alpha,epsilon-dicitraconyl glucagon at pH 4.0 and room temperature regenerated the free epsilon-amino group within 16 h, while the citraconyl-alpha-amino group was stable. N alpha-Citraconyl glucagon was purified by anion-exchange chromatography and was a weak partial agonist in stimulating adenylate cyclase in rat liver plasma membranes. The derivative exhibited 1% of the biological potency and 35-40% of the maximal stimulation of glucagon. Binding affinity to plasma membranes was also reduced, but not to as great an extent as adenylate cyclase activity. Removal of the alpha-citraconyl group by treatment with 10 mM HCl at 40 degrees C restored full potency and stimulation to glucagon. These results suggest that the N-terminal histidine of glucagon is involved in both binding to plasma membranes and transduction of the signal to adenylate cyclase.
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D001681:Biological Assay; D002462:Cell Membrane / Q000378:metabolism; D004789:Enzyme Activation; D005934:Glucagon / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism; D018027:Receptors, Glucagon
D011956:Receptors, Cell Surface; D018027:Receptors, Glucagon; C036349:glucagon, N(alpha)-citraconyl; D005934:Glucagon; D000262:Adenylyl Cyclases