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.
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 intramolecular high-mannose chains in the folding and assembly of soybean lectin polypeptides, the effects of asparagine-linked oligosaccharides, Man9GlcNAc2Asn (M9-Asn) and Glc1-3Man9GlcNAc2Asn (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 300 microM 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 Man alpha 1-2Man-alpha 1-2Man linked to the 3 position of the beta-mannosyl residue of the high-mannose chains functions in the folding of the subunit polypeptides, and that other branches participate in the subunit assembly.
Nagai
K
K
Department of Agricultural Chemistry, College of Agriculture, University of Osaka Prefecture.
Shibata
K
K
Yamaguchi
H
H
D001216:Asparagine / Q000737:chemistry; D002236:Carbohydrate Conformation / Q000187:drug effects; D002240:Carbohydrate Sequence; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D037102:Lectins / Q000096:biosynthesis / Q000737:chemistry* / Q000187:drug effects; D046911:Macromolecular Substances; D008358:Mannose / Q000737:chemistry*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis / Q000737:chemistry*; D010452:Peptide Biosynthesis; D010455:Peptides / Q000737:chemistry*; D037121:Plant Lectins; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation; D017510:Protein Folding; D030262:Soybean Proteins; D013025:Glycine max / Q000737:chemistry*; D010835:Phytohemagglutinins
D001216:Asparagine; D019791:Guanidine; D006146:Guanidines; D037102:Lectins; D046911:Macromolecular Substances; D008358:Mannose; D009844:Oligosaccharides; D010455:Peptides; D037121:Plant Lectins; D030262:Soybean Proteins; C028824:soybean lectin; D010835:Phytohemagglutinins