title: Freeze tolerance in the wood frog Rana sylvatica is associated with unusual structural features in insulin but not in glucagon.
abstract: The wood frog Rana sylvatica utilises glucose, derived from hepatic glycogen, as a cryoprotectant in order to survive freezing during winter hibernation, and glycogenolysis is initiated by hormonal and/or neural stimuli. The primary structure of insulin was determined from R. sylvatica and from two species of freeze-intolerant Ranid frogs R. catesbeiana (American bullfrog) and R. ridibunda (European green frog). All three insulins contain a dipeptide (Lys-Pro) extension to the N-terminus of the A-chain. The amino acid sequences of insulins from R. catesbeiana and R. ridibunda differ by only one residue (Asp for Glu at B21) but R. sylvatica insulin differs from R. catesbeiana insulin at A12 (Thr-->Met), A23 (Asn-->Ser), B5 (Tyr-->His) and B13 (Glu-->Asp). The residue at A23 (corresponding to A21 in human insulin) has been otherwise fully conserved during evolution and the residue at B13 has been strongly conserved in tetrapods. Insulin isolated from specimens of R. sylvatica that had been frozen for 24 h and from control animals that had not been frozen had the same structure, showing that freezing did not alter the pathway of post-translational processing of proinsulin. R. sylvatica glucagon was isolated in two molecular forms. Glucagon-29 was identical to R. catesbeiana glucagon-29 and contains only one amino acid substitution (Thr-->Ser) compared with human glucagon. Glucagon-36 represents glucagon-29 extended from its C-terminus by Lys-Arg-Ser-Gly-Gly-Ile-Ser and is identical to R. catesbeiana glucagon-36. We speculate that selective changes in the structure of the insulin molecule may contribute to the anomalous regulation of glycogen phosphorylase in the wood frog.
authors.affiliation: Regulatory Peptide Center, Department of Biomedical Sciences, Creighton University School of Medicine, Omaha, Nebraska 68178-0405, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Conlon
authors.forename: K
authors.initials: K
authors.lastname: Yano
authors.forename: N
authors.initials: N
authors.lastname: Chartrel
authors.forename: H
authors.initials: H
authors.lastname: Vaudry
authors.forename: K B
authors.initials: KB
authors.lastname: Storey
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005615:Freezing; D005934:Glucagon / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D007328:Insulin / Q000737:chemistry* / Q000302:isolation & purification; D008297:Male; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011898:Ranidae / Q000502:physiology*; D017386:Sequence Homology, Amino Acid
chemical_list: D007328:Insulin; D005934:Glucagon