TY - JOUR
T1 - Purification of Rat Liver Fucose Binding Protein
AU - Lehrman, Mark A.
AU - Hill, Robert L.
N1 - Funding Information:
This work was supported by the National Institutes of Health, Institute of General Medical Sciences, Grant GM-2747.
PY - 1983/1/1
Y1 - 1983/1/1
N2 - 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.
AB - 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.
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U2 - 10.1016/0076-6879(83)98159-4
DO - 10.1016/0076-6879(83)98159-4
M3 - Article
C2 - 6669053
AN - SCOPUS:0021007153
SN - 0076-6879
VL - 98
SP - 309
EP - 320
JO - Methods in Enzymology
JF - Methods in Enzymology
IS - C
ER -