DEMONSTRATION OF PERIPHERAL FUCOSE UNITS IN N-LINKED GLYCANS OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 GP-120 - EFFECTS ON GLYCOPROTEIN CONFORMATION

Fucosylated N-linked glycans are important constituents of membrane gl ycoproteins, owing to their significance as biologically active ligand s for several selectins and their role in modulating protein conformat ion of viral glycoproteins. The human immunodeficiency virus type 1 (H IV-1) glycoprotein contains more than 30 different glycan structures b ut so far fucose was found associated solely with the innermost GlcNAc of N-linked glycans. In the present report we determined whether fuco se units also were linked to the distal GlcNAc via alpha(1-3) or alpha (1-4) linkages in N-linked glycans of gp 120. [H-3]-fucose labelled gp 120 was subjected to endoglycosidase F digestion, releasing diantenna ry complex type N-linked glycans, but leaving the inner polypeptide-bo und carbohydrates, GlcNAc and possibly associated fucose units, intact . Gel filtration of the digested material revealed that [H-3]-fucose l abel was released from gp 120 by this treatment, indicating presence o f peripheral fucose units. Furthermore, [H-3]-focuse label was also re leased by treatment of the labelled gp 120 with an alpha-1-fucosidase specifically removing fucose in alpha(1-3) and alpha(1-4) linkages. Al together the results indicated presence of fucose units linked to peri pheral GlcNAc of gp 120 N-linked glycans. We have earlier shown that o ther peripheral carbohydrate determinants, i.e. beta(1-4)galactose on N-linked glycans, maintain a correct antigenic conformation of gp 120. Using a coupled ELISA system, where changes in antigenic behaviour of a viral glycoprotein were correlated to stepwise elimination of perip heral monosaccharides from N-linked glycans, we found that treatment o f gp 120 with a pan-specific alpha-fucosidase as well as an enzyme spe cific for alpha(1-3)- or alpha(1-4)-linked fucose disclosed a hidden l inear epitope situated in the gp 120 C2 region. The effects of the gen eral fucosidase on epitope exposure was more prominent than those obta ined with the enzyme with narrow specificity, suggesting that peripher al and inner fucose units co-operate in the maintenance of gp 120 conf ormation.