Tyrosine-272 is involved in the inhibition of protein phosphatase-1 by multiple toxins.
Protein phosphatase-1 (PP1) is regulated by interaction with different subunits, which include several inhibitory proteins. It is also potently inhibited by several toxins of diverse origins. Recent work has identified a region near the C-terminus of PP1 (residues 274-277) whose modification was shown to moderate okadaic acid binding [Zhang et al. (1994) J. Biol. Chem. 269, 16997-17000]. In this study, the role of this region in toxin binding was explored by site-directed mutagenesis. A residue (Tyr-272) was identified whose mutation had dramatic effects on the spectrum of inhibitor sensitivity of PP1. The IC50's of a number of mutants of Tyr-272 toward okadaic acid, tautomycin, calyculin A, microcystin-LR, nodularin, inhibitor-2, and cantharidic acid were determined and compared to that of the wild-type enzyme. The sensitivity of PP1 toward tautomycin and calyculin A was markedly decreased, by as much as 3 orders of magnitude, with lesser effects on okadaic acid and nodularin, and with microcystin-LR and inhibitor-2 being the least affected. These studies show that Tyr-272 is of specific importance for the binding of these inhibitors and provide strong evidence for the postulate that these toxins all bind to a common inhibitor site on PP1. In addition, our studies show that Tyr-272 is not required for catalytic activity.
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Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, Florida 33101, USA.
Zhang
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Long
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Lee
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D000595:Amino Acid Sequence; D003545:Cysteine; D004252:DNA Mutational Analysis; D004351:Drug Resistance; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004988:Ethers, Cyclic / Q000494:pharmacology; D008969:Molecular Sequence Data; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D054645:Protein Phosphatase 1; D013329:Structure-Activity Relationship; D014118:Toxins, Biological / Q000494:pharmacology*; D014443:Tyrosine
D004791:Enzyme Inhibitors; D004988:Ethers, Cyclic; D014118:Toxins, Biological; D019319:Okadaic Acid; D014443:Tyrosine; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D003545:Cysteine