Isolation of collagen from the skins of Ehlers-Danlos syndrome-affected dogs by acetic acid extraction and pepsin digestion.
Collagen was isolated by acetic acid extraction in the presence of protease inhibitors and also by pepsin digestion from the skins of dogs affected with the Ehlers-Danlos syndrome and the skins on non-affected dogs. The collagen preparations isolated by acetic acid extraction from the Ehlers-Danlos syndrome-affected dog skin contained a greater proportion of alpha-chains than the collagen preparations from the normal dog skin. When the collagen from the Ehlers-Danlos syndrome-affected dog skin was reduced with NaBH4 before heat denaturation, and electrophoresis, there was a greater proportion of beta-chains present. The collagen isolated from the normal dog skin was not affected by the NaBH4 reduction. Collagen preparations isolated by pepsin digestion from both the Ehlers-Danlos syndrome-affected dog skin and the non-affected dog skin contained the same quantity of alpha- and beta-chains. In addition, collagen from both affected and non-affected dog skins isolated by pepsin digestion contained 10-11% type III collagen as determined by the interrupted sodium dodecyl sulfate polyacrylamide gel electrophoresis method. Pepsin digestion of the collagens isolated by acetic acid extraction in the presence of protease inhibitors from the skins of affected and non-affected dogs eliminated the differences between the alpha:beta ratios of the affected and non-affected collagen preparations.
D000085:Acetates; D000818:Animals; D001894:Borohydrides; D003094:Collagen / Q000302:isolation & purification*; D004283:Dog Diseases / Q000378:metabolism*; D004285:Dogs; D004535:Ehlers-Danlos Syndrome / Q000378:metabolism / Q000662:veterinary*; D046911:Macromolecular Substances; D010084:Oxidation-Reduction; D010434:Pepsin A; D012867:Skin / Q000032:analysis*
D000085:Acetates; D001894:Borohydrides; D046911:Macromolecular Substances; D003094:Collagen; D010434:Pepsin A