Neutral proteinases of the human intervertebral disc.
Disc tissue consisting of pooled annuli fibrosus and nuclei pulposus from the cadaver of an adolescent aged 19 years was extracted with 4.0 M Gu-HCl. Proteins of low buoyant density (p less than or equal to 1.38 g/ml) containing the disc enzymes and inhibitors were separated from proteoglycans of high buoyant density (p greater than or equal to 1.50 g/ml) by density gradient ultracentrifugation. Sephadex G-75F gel chromatography followed by trypsin affinity chromatography was then used to resolve disc proteolytic and trypsin inhibitory activities. The results obtained were strongly suggestive of the presence of a high molecular weight zymogen which upon activation generated a population of smaller molecular weight proteinases. The disc proteinases obtained by this process showed similar properties in terms of: their pH optima (7.4-7.6); their inhibition patterns by class-specific proteinase inhibitors; their variation of activity as a function of NaCl and lysine concentrations; and the hydrodynamic size of their proteoglycan degradation products. The activated disc neutral proteinase demonstrated many characteristics in common with plasmin; however, unlike the latter, the disc proteinases also showed some calcium dependence.
D000293:Adolescent; D000328:Adult; D002845:Chromatography; D002850:Chromatography, Gel; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D007403:Intervertebral Disc / Q000201:enzymology*; D007976:Leupeptins / Q000494:pharmacology; D008297:Male; D010662:Phenylmercuric Acetate / Q000031:analogs & derivatives / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D014461:Ultracentrifugation
D007976:Leupeptins; D011480:Protease Inhibitors; D012965:Sodium Chloride; C010024:4-aminophenylmercuriacetate; D010450:Endopeptidases; D010662:Phenylmercuric Acetate