CHARACTERIZATION OF DELETION MUTANTS OF THE CATALYTIC SUBUNIT OF PROTEIN PHOSPHATASE-1

Deletion mutagenesis was used to define the core region of the catalyt ic subunit of rabbit muscle protein phosphatase-1. Deletions in the N terminus were found to lead to loss of expression. Deletions of up to 33 residues from the C-terminal region were tolerated, and the truncat ed enzymes were fully active. Deletion of an additional 21 residues le d to loss of expression. Mutants which had had 33 and 25 residues dele ted maintained specific activities that were comparable to those of th e wild type enzyme. The response of these two deletion mutants to okad aic acid, microcystin, and inhibitor-2 was determined, Only slightly l ower IC50 values were observed in all cases, showing that the C termin us itself does not play a major role in the binding of these inhibitor s. The deletion mutants formed stable complexes with inhibitor a as sh own by gel filtration. These studies provide unambiguous evidence that the extreme C-terminal region of protein phosphatase-1 is not directl y involved in catalytic function or in the binding of inhibitor-2, mic rocystin, or okadaic acid, and they also establish that the first simi lar to 300 residues of the sequence constitute a sufficient core for p rotein phosphatase-1 catalytic functions.