Characterization of an insulin from the three-toed amphiuma (Amphibia: Urodela) with an N-terminally extended A-chain and high receptor-binding affinity Available to Purchase

Insulin was isolated from an extract of the pancreas of a urodele, the three-toed amphiuma (Amphiuma tridactylum), and its primary structure established as Ala-Arg-Gly-Ile-Val-Glu-Gln-Cys-Cys-His¹⁰-Asn-Thr-Cys-Ser-Leu-Asn-Gln-Leu-Glu-Asn²⁰-Tyr-Cys-Asn for the A-chain and Ile-Thr-Asn-Gln-Tyr-Leu-Cys-Gly-Ser-His¹⁰-Leu-Val-Glu-Ala-Leu-Tyr-Leu-Val-Cys-Gly²⁰-Asp-Arg-Gly-Phe-Phe-Tyr-Ser-Pro-Lysfor the B-chain. The N-terminus of the A-chain is extended by two amino acids (Ala-Arg) relative to all other known insulins suggesting an anomalous pathway of post-translational processing in the region of the C-peptide/A-chain junction of proinsulin. In common with chicken and Xenopus insulins, which contain a His^A8, amphiuma insulin was more potent (approx. 5-fold) than porcine insulin in inhibiting the binding of [¹²⁵I-Tyr^A14]insulin to the soluble human insulin receptor from transfected 293EBNA cells (an adenovirus-transformed human kidney cell line). This result is consistent with previous data showing that insulin analogues extended at Gly^A1 by uncharged groups have reduced binding affinity whereas high affinity is preserved in analogues extended by basic amino acid residues.