Role of Intramolecular High-Mannose Chains in the Folding and Assembly of Soybean (Glycine max) Lectin Polypeptides: Studies by the Combined Use of Spectroscopy and Gel-Filtration Size Analysis

Abstract

It was previously reported [Nagai, K. & Yamaguchi, H. (1993) J. Biochem. 113, 123-125] that intramolecular high-mannose chains are essential for reconstitution of soybean lectin from denatured subunits. To obtain more detailed information on the role of the intra-molecular high-mannose chains in the folding and assembly of soybean lectin polypep-tides, the effects of asparagine-linked oligosaccharides, Man₉GIcNAc₂Asn (M9-Asn) and Glc_1-3Man₉GlcNAc₂Asn (GM9-Asn), on the reconstitution of soybean lectin from denatured subunits were examined by comparison with the denaturation features of the lectin with varying concentrations of guanidine hydrochloride. The combined use of spectroscopy and size-analysis by gel filtration revealed that both the folding and assembly of denatured subunit polypeptides were completely prevented in the presence of 300μM M9-Asn, whereas the same concentration of GM9-Asn only interfered with the polypeptide assembly, exhibiting no significant effect on the polypeptide folding. These results, considered together with those in the previous report, indicate that the sugar branch Manα1-2Man-α1-2Man linked to the 3 position of the β-mannosyl residue of the high-mannose chains functions in the folding of the subunit polypeptides, and that other branches participate in the subunit assembly.