This chapter describes the preparation of a liver lectin that has a high affinity for fucose-containing ligands. The starting material for preparation of the fucose binding protein (F-BP) is a Triton X-100 extract of rat liver, which contains F-BP in addition to the galactose (G-BP) and mannose/N-acetylglucosamine binding proteins (M-BP). These methods permit the resolution of the different lectins from one another and further purification of each of the different lectins. Moreover, in developing these methods, a form of the M-BP with 10–20 times the specific activity of most of the M-BP in liver extracts was found to be a major contaminant of the F-BP; methods for preparation of this form of M-BP, designated HM-BP, are also presented in this chapter. Affinity chromatography on columns of the neoglycoprotein L-fucosylbovine serum albumin (Fuc-BSA) is the principal method for purifying the rat liver lectins. F-BP, M-BP, and HM-BP are adsorbed to Fuc-BSA-agarose whereas G-BP is not adsorbed. The bulk of the M-BP is subsequently removed from F-BP by rechromatography on Fuc-BSA-agarose in the presence of N-acetylglucosamine, which inhibits adsorption of M-BP, but not of F-BP or HM-BP.
ASJC Scopus subject areas
- Molecular Biology