Purification, characterization, and biological activity of insulins from the spotted dogfish, Scyliorhinus canicula, and the hammerhead shark, Sphyrna lewini.
Insulin was purified from pancreatic extracts of two elasmobranch species belonging to different families in the order Carcharhiniformes, the European spotted dogfish, Scyliorhinus canicula (Scyliorhinidae), and the hammerhead shark, Sphyrna lewini (Carcharhinidae). The amino acid sequence of dogfish insulin was established as A-chain GIVDHCCRNT(10)CSLYDLEGYC(20)NQ and B-chain LPSQHLCGSH(10)LVETLYFVCG(20)QKGFYYVPKV(30). The primary structure of hammerhead shark insulin was similar to that of dogfish insulin with only 2 amino acid substitutions at A8 (R --> H) and B30 (V --> I). The elasmobranch insulins were markedly different from human insulin (17 amino acid substitutions) but all the residues in human insulin that are believed to be important in determining the receptor binding conformation (B6, B8, B11, B13, B23, B24, B25, A2, A3, and A19) have been conserved in the elasmobranch insulins with the exception of the conservative substitution Phe --> Tyr at B25. Consistent with this, dogfish and human insulin showed almost identical binding affinity to the recombinant solubilized human insulin receptor (K(D) values of 14.0 and 18.6 pM, respectively; relative potency 133%). Previous studies have shown that bovine insulin produces severe and sustained hypoglycemia in elasmobranchs but the effect is of slow onset. Bolus arterial injections of dogfish insulin (10 nmol x kg(-1)) into unanesthetized, fasting dogfish (n = 9) produced no changes in blood glucose, 3-hydroxybutyrate, and acetoacetate concentrations over a 4-h period. In a second series of experiments (n = 7), dogfish insulin (10 nmol x kg(-1)) produced a significant (P < 0.05) fall in blood glucose after 12 h that persisted for at least 48 h, but no change in ketone body concentrations. The data indicate that the metabolic actions of an endogenous elasmobranch insulin in an elasmobranch are similar to those previously described for mammalian insulin.
Anderson
W Gary
WG
Gatty Marine Laboratory, School of Biology, St Andrews KY16 8LB, United Kingdom.
Ali
Mohamed F
MF
Einarsdóttir
Ingibjörg E
IE
Schäffer
Lauge
L
Hazon
Neil
N
Conlon
J Michael
JM
D020155:3-Hydroxybutyric Acid / Q000097:blood; D000090:Acetoacetates / Q000097:blood; D000595:Amino Acid Sequence; D000818:Animals; D001786:Blood Glucose / Q000032:analysis / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000662:veterinary; D017124:Conserved Sequence; D004284:Dogfish / Q000378:metabolism* / Q000502:physiology; D005260:Female; D006801:Humans; D007328:Insulin / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D011972:Receptor, Insulin / Q000378:metabolism
D000090:Acetoacetates; D001786:Blood Glucose; D007328:Insulin; C016635:acetoacetic acid; D011972:Receptor, Insulin; D020155:3-Hydroxybutyric Acid