title: Glucagon carboxyl-terminal derivatives: preparation, purification, and characterization.
abstract: Chemical and enzymatic methods have been used to prepare the following series of seven glucagon derivatives modified in the carboxyl-terminal region important for hormone-receptor binding: [des-Asn28,Thr29](homoserine lactone27)glucagon, [des-Asn28,Thr29](homoserine27)glucagon, (S-methyl-Met27)glucagon, [des-Thr29](S-methyl-Met27)-glucagon, [des-Thr29]glucagon,[des-Asn28,Thr29](S-methyl-Met27)glucagon, and [des-Asn28,Thr29]glucagon. The derivatives were isolated in high yield, extensively purified, and chemically characterized. All were found to be full agonists of native glucagon. Binding affinity was evaluated by displacement of mono[125I]iodoglucagon prepared by new methods. Binding and biological activities closely correlated, indicating that most modifications affected the relative binding affinity and relative biological potency of glucagon to a comparable extent. Circular dichroism measured in dilute acid solution resembled that of native glucagon except for [des-Asn28,Thr29]glucagon which displayed increased alpha helicity (25%). All derivatives formed helical structures in 2-chloro-ethanol, although the amount of helicity induced was not closely correlated with biological activity. Binding and biological activities were not affected by removal of Thr-29, though both were reduced 20-fold when Asn-28 was also removed, irrespective of whether homoserine or native methionine remained at the carboxyl terminus. Lactone formation was associated with a further 5-fold reduction in binding affinity but not in activity. Methylation of Met-27 had essentially the same effect as removing the two carboxyl-terminal residues, although the combined effect of both modifications was greater than 100-fold reduction in binding and activity. These findings provide additional insight concerning glucagon structure-function relationships.
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002268:Carboxypeptidases / Q000378:metabolism; D003488:Cyanogen Bromide; D004789:Enzyme Activation / Q000187:drug effects; D005934:Glucagon / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000494:pharmacology; D007467:Iodoproteins; D008099:Liver / Q000201:enzymology; D008745:Methylation; D010446:Peptide Fragments; D051381:Rats; D013329:Structure-Activity Relationship
chemical_list: D007467:Iodoproteins; D010446:Peptide Fragments; D005934:Glucagon; D002268:Carboxypeptidases; D000262:Adenylyl Cyclases; D003488:Cyanogen Bromide