Hydroxyapatite Induces Autolytic Degradation and Inactivation of Matrix Metalloproteinase-1 and -3

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Abstract
In the course of studies to identify a protease capable of producing a long‐lived 50 kDa fragment of bone acidic glycoprotein‐75 (BAG‐75), it was observed that incubation of matrix metalloproteinase (MMP)‐3 (stromelysin 1) with preparations of BAG‐75 led to inactivation of proteolytic function, e.g., an inability to fragment125I‐labeled BAG‐75 added subsequently. MMP‐1 (interstitial collagenase) was also inactivated by exposure to BAG‐75 preparations. Investigation of the mechanism revealed that BAG‐75 preparations contained millimolar levels of inorganic phosphate which formed hydroxyapatite crystals under digestion conditions. Hydroxyapatite crystals alone and in BAG‐75–hydroxyapatite complexes induced the autolytic degradation of both active and precursor forms of MMP‐1 and MMP‐3. Autolytic degradation in the presence of hydroxyapatite was demonstrated by a loss in catalytic function assayed with peptide and/or protein substrates, and, by fragmentation into polypeptides of 1000 μg/ml), suggesting that autolysis may be mediated by bound enzyme. The effect of hydroxyapatite appears to be specific for MMP‐1 and MMP‐3 since the catalytic activity of chymotrypsin, trypsin, papain, and thermolysin remained unchanged after exposure to hydroxyapatite. These results document for the first time a novel catalytic role for hydroxyapatite crystals in vitro and provide an initial biochemical characterization of the intermolecular, autolytic, calcium ion‐dependent, matrix metalloproteinase‐specific degradative mechanism.