Purification and characterization of an acid metalloproteinase from human articular cartilage.
A metalloprotease that digests cartilage proteoglycan optimally at pH 5.3 has been purified (4400-fold) to homogeneity from 20-g samples of human articular cartilage containing about 100 micrograms of enzyme. This enzyme was cleanly separated from a related neutral metalloprotease with an optimum pH of 7.2. The acid metalloprotease displays 40% of its maximum activity at pH 7.2 and so has significant activity at physiological pH. The protease is calcium-dependent and indirect evidence suggests that it may contain zinc at its active center. It occurs largely in a latent form that can be activated by aminophenylmercuric acetate. The apparent Mr of the latent form is 55,000 and of the active form, 35,000. The isoelectric point is at pH 4.9. The protease activity is inhibited by chelators, Z-phenylalanine, ovostatin, and tissue inhibitor of metalloproteinase from human articular cartilage. It differs from metalloproteinases such as enkephalinase and kidney brush-border protease in its failure to be strongly inhibited by phosphoramidon and Zincov. It cleaves the proteoglycan monomer of bovine nasal cartilage to fragments of approximately 140,000 Da. It cleaves the B chain of insulin at Ala14-Leu15 and Tyr16-Leu17. A survey of 26 cartilage extracts indicates this enzyme is elevated to about 3 times the normal level in human osteoarthritic cartilage and that the tissue inhibitor of metalloproteinase is only slightly diminished. Preliminary evidence points to the presence of a similar acid metalloprotease activity in human polymorphonuclear leukocytes.
D000595:Amino Acid Sequence; D002358:Cartilage, Articular / Q000201:enzymology*; D002850:Chromatography, Gel; D004527:Egg Proteins / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000302:isolation & purification*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006358:Hot Temperature; D006801:Humans; D007328:Insulin / Q000378:metabolism; D008259:Macroglobulins; D008666:Metalloendopeptidases; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology; D010003:Osteoarthritis / Q000378:metabolism; D010649:Phenylalanine / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011489:Protein Denaturation; D019714:Tissue Inhibitor of Metalloproteinases; D000511:alpha-Macroglobulins
D004527:Egg Proteins; D004791:Enzyme Inhibitors; D007328:Insulin; D008259:Macroglobulins; D011480:Protease Inhibitors; D019714:Tissue Inhibitor of Metalloproteinases; D000511:alpha-Macroglobulins; D010649:Phenylalanine; C036828:ovomacroglobulin; D010450:Endopeptidases; D008666:Metalloendopeptidases