Selective solubilization of two populations of polypeptides from bovine retinal basement membranes.
Basement membranes isolated from the entire neural retinae of normal cows consist of a heterogeneous mixture of polypeptides resolvable as two major populations, one collagenous and the other non-collagenous. The non-collagenous population consists of disulfide-bonded aggregates of which the major fraction is soluble in lithium dodecyl sulfate (LDS) at 4 degrees C without reducing agent, with the remainder requiring reduction for solubilization at 4 degrees C. Only the second 4 degrees C extract is contaminated with small amounts of hydroxylated amino acids. The non-collagenous population consists of up to 25 polypeptides detectable in sodium dodecyl sulfate (SDS) gels, all of which stain blue with Coomassie Blue R-250, are insensitive to bacterial collagenase and migrate on or slightly below the central diagonal in urea-SDS/SDS electrophoresis. The collagenous population is soluble in LDS at 100 degrees C under reducing conditions and has a collagenous amino acid composition, although consisting of only 272 glycine residues/1000. This fraction consists of four major and four minor polypeptides detectable in SDS gels, all of which stain red (metachromatically) with Coomassie Blue R-250, are degraded by bacterial collagenase and migrate below the central diagonal in urea-SDS/SDS electrophoresis. The four major collagenous polypeptides are larger than the alpha-chains of type I collagen. In total, the solubilized proteins account for 60% of the general protein, but only 30% of the sum of hydroxyproline and hydroxylysine. The insoluble residue has a collagenous amino acid composition similar to that of the 100 degrees C extract, but with lower 3-hydroxyproline and hydroxylysine contents.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001485:Basement Membrane / Q000032:analysis / Q000648:ultrastructure; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D005233:Fatty Alcohols; D003012:Microbial Collagenase; D008854:Microscopy, Electron; D010455:Peptides / Q000302:isolation & purification*; D012160:Retina / Q000032:analysis* / Q000648:ultrastructure; D012967:Sodium Dodecyl Sulfate; D012995:Solubility
D000596:Amino Acids; D005233:Fatty Alcohols; D010455:Peptides; D012967:Sodium Dodecyl Sulfate; C028913:dodecyl sulfate; D003012:Microbial Collagenase