Semisynthetic derivatives of glucagon. The contribution of histidine-1 to hormone conformation and activity.
Semisynthetic N epsilon- acetimidoglucagon was prepared from the [des- His1 ]analogue by coupling the N-hydroxysuccinimide ester of N alpha- tBoc - Nimidazole -DNP-L-histidine to the peptide in dimethylformamide in the presence of 1-hydroxybenzotriazole. The deprotected, purified product was chemically identical to N epsilon- acetimidoglucagon and equipotent to N epsilon- acetimidoglucagon and native glucagon in its ability to activate adenylate cyclase and displace [125I] iodoglucagon from rat liver plasma membranes. Semisynthetic [ Phe1 ]-, [ Ala1 ]-, and [des- His1 ] glucagons prepared similarly achieved 85, 55, and 35% of the maximal activity and 22, 2, and 6% of the binding potency of N epsilon- acetimidoglucagon . The biological assays indicate that the amino group is involved to a greater extent in transduction than in binding, but the aromatic nature and hydrogen bonding capability of the imidazole ring of histidine-1 are important for both binding and transduction. In circular dichroism studies, all derivatives exhibited increased helicity in 2-chloroethanol. The [ Phe1 ] analogue although less soluble behaved similarly to native glucagon, while the [ Ala1 ] and [des- His1 ] derivatives exhibited an increased helical content in 0.01 N HCl as a result of an increased propensity of these derivatives to self-associate in the absence of 2-chloroethanol. The unexpected conformational changes throughout the molecule may have relevance for the functional activity.
D000262:Adenylyl Cyclases / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002942:Circular Dichroism; D005934:Glucagon / Q000031:analogs & derivatives* / Q000378:metabolism; D006639:Histidine; D007525:Isoelectric Focusing; D008099:Liver / Q000201:enzymology; D011487:Protein Conformation; D051381:Rats
D000596:Amino Acids; D006639:Histidine; C041452:N(epsilon)-acetimidoglucagon; C036044:N(epsilon)-acetimidoglucagon, des-His(1)-; D005934:Glucagon; C041454:N(epsilon)-acetimidoglucagon, Phe(1)-; C041453:N(epsilon)-acetimidoglucagon, Ala(1)-; D000262:Adenylyl Cyclases