In vitro characterization and in vivo clearance of recombinant barramundi (Lates calcarifer) IGF-I

Little is known about fish insulin-like growth factors (IGFs) as only small amounts have been isolated from native sources or indeed produced recombin antly. This report describes the production of milligram quantities of reco mbinant barramundi IGF-I (bIGF-I) and its subsequent characterization. Reco mbinant bIGF-I was produced in Escherichia coli using a gene fusion system similar to that previously described for the production of other non-mammal ian IGFs. Recombinant bIGF-I was similar to human IGF-I (hIGF-I) in stimula ting protein synthesis and in competing for binding of labelled hIGF-I to I GF receptors whether tested in rat myoblasts or in salmon embryo fibroblast s. However, recombinant bIGF-I differed from its human counterpart in its a ffinity for a polyclonal antibody raised against hIGF-I, with at least 200- fold more bIGF-I required to obtain 50% displacement of labelled hIGF-I fro m the antibody. Hence, the recombinant protein will be essential for develo ping a specific homologous immunoassay for measuring IGF-I concentrations i n barramundi during growth and development. In addition, studies investigat ing the clearance of labelled bIGF-I and hIGF-I in vivo reveal that the hum an protein is cleared from the circulation of juvenile barramundi almost tw ice as fast as the barramundi protein, thus providing the first in vivo evi dence that there are functional differences between fish and human IGF-Is. Neutral gel chromatography of serum from the clearance study suggest that t his is due to differences in the affinities of the labelled human and fish IGF-I for the IGFBPs present in barramundi. (C) 1999 Elsevier Science B.V. All rights reserved.