Separation and identification of carp pituitary proteins and glycoproteins

Carp pituitary proteins and glycoproteins were separated by the combination of immobilized metal affinity chromatography (IMAC) and affinity chromatography on Con A-Sepharose. The protein fractions were analysed by SDS-PAGE. Luteinizing hormone (in the alpha(1)beta and alpha(2)beta forms), growth hormone, free alpha(1) subunit and beta subunit of thyroid-stimulating hormone were identified by N-terminal amino acid sequencing. N-linked oligosaccharide chains of thyroid-stimulating hormone (beta TSH) were separated after fluorescent labelling on a GlycoSep N column and treated by exoglycosidases. Among the saccharide components, complex and hybrid structures terminating SO4-GalNAc-GlcNAc-Man-, and high mannose structures, with 1 to 8 mannose units attached to the oligosaccharide core (GlcNAc)(2)(Man)(3), were found.