Structural and biochemical characterization of inhibitor-1α

Inhibitor-1α is one of the isoforms of human protein phosphatase inhibitor-1. It is a product of alternative splicing of inhibitor-1 gene and lacks 51 internal amino acids from residue 84 to 134 of inhibitor-1. Here we have characterized the structural and biochemical properties of inhibitor-1α. Structural analysis of recombinant inhibitor-1α by NMR spectroscopy revealed that inhibitor-1α adopts a predominantly random coil conformation. Excluding the region from residue 84 to 134 of inhibitor-1, the structural features of inhibitor-1 and inhibitor-1α are almost the same as each other. The IC₅₀ value of inhibitor-1α in inhibition of Protein phosphatase-1 (PP1) is comparable to that of inhibitor-1, indicating that inhibitor-1α is a potent inhibitor of PP1 when Thr-35 is phosphorylated by PKA. For phosphorylation by PKA and dephosphorylation by protein phosphatase-1, -2A, and -2B, the measured kinetic parameters of inhibitor-1α are very close to those of inhibitor-1. Taken together, these results suggest that inhibitor-1α preserves the structure of inhibitor-1, the PP1 inhibitory activity and the functional specificities toward phosphorylation by PKA and dephosphorylation by protein phosphatase-1, -2A, and -2B.