Structural evidence for a functional role of human tissue nonspecific alkaline phosphatase in bone mineralization

The human tissue nonspecific alkaline phosphatase (TNAP) is found in liver, kidney, and bone. Mutations in the TNAP gene can lead to Hypophosphatasia, a rare inborn disease that is characterized by defective bone mineralizati on. TNAP is 74% homologous to human placental alkaline phosphatase (PLAP) w hose crystal structure has been recently determined at atomic resolution (L e Du, M. H., Stigbrand, T., Taussig, M. J., Menez, A., and Stura, E. A. (20 01) J. Biol. Chem, 276,9158-9165). The degree of homology allowed us to bui ld a reliable TNAP model to investigate the relationship between mutations associated with hypophosphatasia and their probable consequences on the act ivity or the structure of the enzyme. The mutations are clustered within fi ve crucial regions, namely the active site and its vicinity, the active sit e valley, the homodimer interface, the crown domain, and the metal-binding site. The crown domain and the metal-binding domain are mammalian-specific and were observed for the first time in the PLAP structure. The crown domai n contains a collagen binding loop. A synchrotron radiation x-ray fluoresce nce study confirms that the metal in the metal-binding site is a calcium io n. Several severe mutations in TNAP occur around this calcium site, suggest ing that calcium may be of critical importance for the TNAP function. The p resence of this extra metal-binding site gives new insights on the controve rsial role observed for calcium.