Purification of a 41-kDa insulin-like growth factor binding protein from serum of chinook salmon, Oncorhynchus tshawytscha

In salmon, at least three insulin-like growth factor binding proteins (IGFBPs) with molecular masses of 41, 28, and 22 kDa exist in serum. The 41-kDa IGFBP is up-regulated by growth hormone treatment and down-regulated by fasting, suggesting that it is a homolog of IGFBP-3. We purified the 41-kDa IGFBP from chinook salmon serum by IGF-I affinity chromatography followed by reversed-phase high pressure liquid chromatography. Purified IGFBP appeared as doublet bands on electrophoresis and was N-glycosylated. Analysis of partial N-terminal amino acid sequence revealed that salmon 41-kDa IGFBP has the cysteine rich domain conserved among IGFBP family. In a binding assay using 125 I-salmon IGF-I, purified 41-kDa IGFBP specifically bound salmon IGF-I, human IGF-I and human IGF-II, but neither Long R 3 IGF-I nor salmon insulin, showing that binding characteristics of the salmon IGFBP are similar to those of mammalian IGFBPs. Although the partial amino acid sequence of 41-kDa IGFBP showed highest homologies with zebrafish and seabream IGFBP-2, the highly conserved nature of the N-terminus makes it impossible to identify the type of IGFBP from partial sequence data. However, based on physiological responses, molecular weight and type of glycosylation, the 41-kDa IGFBP is most similar to mammalian IGFBP-3.