Isolation and characterization of two different insulins from an amphibian, Xenopus laevis

The South African clawed toad, Xenopus laevis, is a versatile laboratory model of vertebrate development. To study the role of insulin during embryogenesis, we have recently cloned preproinsulin cDNAs from this species. Unexpectedly, we identified two preproinsulin cDNAs corresponding to two different nonallelic genes that code for similar but distinctly different insulins. We now report the isolation, amino acid sequence, and characterization of both of these insulins from pancreatic extracts of adult toads, confirming that both Xenopus preproinsulin genes are expressed. Xenopus insulins represent the first amphibian insulins to be characterized. Xenopus insulin I and Xenopus insulin II are more similar to each other than they are to insulins of other species. In addition, Xenopus insulins are more similar to mammalian and bird insulins, than they are to fish insulins, implying a closer evolutionary link to terrestrial vertebrates than to most aquatic vertebrates. A homogeneous preparation of Xenopus insulin I showed high reactivity in a pork insulin RIA. Xenopus insulin I was approximately 2-fold more potent than pork insulin in binding to insulin receptors on human IM-9 lymphocytes and 1.5-fold more potent than pork insulin in stimulating glucose oxidation in rat adipocytes. We were unable to purify Xenopus insulin II sufficiently for immunological and biological characterization.