Expression of recombinant inhibitor-2 in E. coli and its utilization for the affinity chromatography of protein...

The coding sequence for rabbit muscle protein phosphatase inhibitor-2 was inserted into the pET3a expression vector. This vector allowed the expression of recombinant inhibitor-2 at levels of ca. 1.5% of the soluble protein. A simple procedure allowed the purification of inhibitor-2 from Escherichia coli lysates. This involved heat-treatment, followed by chromatography on Blue-Sepharose and Q-Sepharose. Recombinant inhibitor-2 inhibited rabbit muscle protein phosphatase-1 with a potency similar to that reported for the wild type protein. The recombinant protein was coupled to CH-Sepharose and this support was found to bind the catalytic subunit of protein phosphatase-1 with high efficiency. A procedure for a single-step affinity purification of recombinant ppase-1 from E. coli lysates was shown to be feasible.