A facilitative role for carbonic anhydrase activity in matrix vesicle mineralization.

Carbonic anhydrase (CA) which catalyzes the reversible hydrolysis of carbon dioxide is known to be important in osteoclastic bone resorption, however, suggested roles in calcium phosphate mineral formation have not been previously demonstrated. Biochemical evidence is provided for the presence of CA in growth plate matrix vesicles (MV) and the level of activity determined by enzyme assay. Inhibition of CA activity with the specific inhibitor acetazolamide resulted in reduced rates of MV mineralization. Other inhibitor studies showed that MV mineralization was also impaired by 4,4-diisothiocyanatostilbene-2, 2-disulfonic acid (DIDS), a blocker of membrane bicarbonate channels. No evidence was found for the presence of any proton pumps or channels. When acetazolamide and DIDS were combined, their inhibitory effects on MV mineralization were additive. These findings suggest that MV possess a pH regulation system composed of carbonic anhydrase and a putative bicarbonate channel. This system may function in the MV by providing intraluminal buffering capacity. The control of intravesicular pH is important for the stabilization of the acid-labile nucleational core complex and in preventing the build-up of protons during calcium phosphate phase transformations.