Abstract
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 mineralization. TNAP is 74% homologous to human placental alkaline phosphatase (PLAP) whose crystal structure has been recently determined at atomic resolution (Le Du, M. H., Stigbrand, T., Taussig, M. J., Ménez, A., and Stura, E. A. (2001) J. Biol. Chem, 276, 9158-9165). The degree of homology allowed us to build a reliable TNAP model to investigate the relationship between mutations associated with hypophosphatasia and their probable consequences on the activity or the structure of the enzyme. The mutations are clustered within five crucial regions, namely the active site and its vicinity, the active site 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 domain contains a collagen binding loop. A synchrotron radiation x-ray fluorescence study confirms that the metal in the metal-binding site is a calcium ion. Several severe mutations in TNAP occur around this calcium site, suggesting that calcium may be of critical importance for the TNAP function. The presence of this extra metal-binding site gives new insights on the controversial role observed for calcium.
Highlights
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 mineralization
Its gene is located on chromosome 1p34 –36 [7], and mutations in the TNAP gene have been associated with hypophosphatasia, a rare inherited disorder, characterized by defective bone mineralization
Summary
The human tissue nonspecific alkaline phosphatase (TNAP) is found in liver, kidney, and bone. This difficulty arises mainly from the lack of data concerning the precise role that TNAP plays in bone mineralization This may be partly solved by the use of site-directed mutagenesis of TNAP cDNA and cell transfection to assay residual activity of the mutant AP enzyme [16, 18, 20, 23,24,25]. The model omits a large number of regions of deletions and insertions in TNAP relative to ECAP, in particular concerning the putative functional regions of TNAP Such insertions are present in human placental alkaline phosphatase (PLAP) which is 57% identical to TNAP in sequence and whose crystal structure has been recently solved to 1.8-Å resolution [26]. Few mutations appear more than once as they belong to overlapping domains
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