Revelation of a Catalytic Calcium-Binding Site Elucidates Unusual Metal Dependence of a Human Apyrase

Human soluble calcium-activated nucleotidase 1 (hSCAN-1) represents a new family of apyrase enzymes that catalyze the hydrolysis of nucleotide di- and triphosphates, thereby modulating extracellular purinergic and pyrimidinergic signaling. Among well-characterized phosphoryl transfer enzymes, hSCAN-1 is unique not only in its unusual calcium-dependent activation, but also in its novel phosphate-binding motif. Its catalytic site does not utilize backbone amide groups to bind phosphate, as in the common P-loop, but contains a large cluster of acidic ionizable side chains. By employing a state-of-the-art computational approach, we have revealed a previously uncharacterized catalytic calcium-binding site in hSCAN-1, which elucidates the unusual calcium-dependence of its apyrase activity. In a high-order coordination shell, the newly identified calcium ion organizes the active site residues to mediate nucleotide binding, to orient the nucleophilic water, and to facilitate the phosphoryl transfer reaction. From...