Fractionation of Glycoprotein Components of the Reduced Alkylated Renal Glomerular Basement Membrane

The soluble glycoprotein components of the S-carboxy-methylated glomerular basement membrane were fractionated by gel filtration in sodium dodecyl sulfate on agarose columns of varying porosity and by DEAE-cellulose chromatography in urea. By such techniques, these components, which represented 70 to 80% of the total membrane and ranged in molecular weight from 30,000 to greater than 700,000, were resolved into fractions differing markedly in their amino acid and sugar composition. Some contained large amounts of hydroxyproline, glycine, and hydroxylysine, while others had fewer of these amino acids characteristic of collagen and were richer in amino acids such as aspartic acid, serine, threonine, tyrosine, lysine, histidine, and S-carboxymethyl-cysteine. The fractions with the more collagen-like composition had a higher number of hydroxylysine-linked glucosylgalactose disaccharide units than did the fractions which were more polar in nature, with as many as 35 units per 1000 amino acids occurring in the former and as few as 2 per 1000 amino acids in the latter. The heteropolysaccharide units (consisting of sialic acid, fucose, galactose, mannose, and hexosamines) were more evenly distributed, ranging from 1 to 4 units per 1000 amino acids with a somewhat higher content being present in the more polar fractions. Both the disaccharide and heteropolysaccharide units occurred in components spanning a wide molecular weight range. The former, however, tended to be predominantly associated with high molecular weight material (over 100,000). These results are believed to be consistent with the occurrence in the glomerular basement membrane of both collagen-like and more polar chains, with the former containing both di- and heteropolysaccharide units and the latter having only the polysaccharides. Although disulfide bonds appear to serve as a major crosslink between the peptide chains of this membrane, NH2-terminal amino acid analyses of higher molecular weight fractions suggested that other as yet undefined cross-links may exist.