A MUTANT OF PROTEIN PHOSPHATASE-1 THAT EXHIBITS ALTERED TOXIN SENSITIVITY

A chimeric mutant was constructed in which a 4-amino acid region (GEFD , residues 274-277) of rabbit muscle protein phosphatase-1 was replace d with the sequence YRCG corresponding to residues 267-270 of rabbit p rotein phosphatase-2A. This was based on the finding of a gene mutatio n in okadaic acid-resistant cells which results in a Cys --> Gly conve rsion in protein phosphatase-2A. The YRCG mutant of protein phosphatas e-1 was expressed and purified. The properties of the mutant enzyme we re investigated in terms of its sensitivity toward several toxin inhib itors (okadaic acid, microcystin, nodularin, calyculin A, and canthari dic acid), as well as inhibitor-2. The mutant enzyme exhibited a gain of function in the form of a 10-fold increased sensitivity toward okad aic acid that suggests this region is involved in toxin binding. Signi ficant changes in sensitivity to inhibitor-2 and several of the other toxins were also observed, indicating that these may have a common bin ding region.