Mutation of the toxin binding site of PP-1c: comparison with PP-2B.
The catalytic cores of PP-1c and PP-2B (calcineurin) are structurally conserved. However, PP-2B is resistant to inhibition by toxins of the okadaic acid and cyclic peptide classes, while PP-1c is potently inhibited. Molecular docking of the structure of microcystin-LR onto the catalytic core of PP-2B identified residues that may be responsible for blocking access of toxins to the catalytic site. Amino acids in PP-1c were substituted with these PP-2B residues to investigate their contribution to PP-2B toxin resistance. Mutants of PP-1c were also produced to test the importance of hydrophobic interactions to toxin binding. Our results suggest that different classes of toxin inhibitors interact with the same hydrophobic side chains of PP-1c through different mechanisms. Substitution of amino acids in PP-1c with PP-2B residues demonstrated no highly significant changes in toxin inhibition. We hypothesize that an interaction outside the catalytic core causing the L7 loop of PP-2B to block the catalytic site may be responsible for PP-2B resistance to toxins.
Dawson
J F
JF
Department of Biochemistry, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada. jf_dawson@hotmail.com
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Bagu
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Holmes
C F
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D000818:Animals; D001665:Binding Sites; D019703:Calcineurin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D002417:Cattle; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008387:Marine Toxins; D052998:Microcystins; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D019319:Okadaic Acid / Q000378:metabolism; D010456:Peptides, Cyclic / Q000378:metabolism; D011487:Protein Conformation; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014118:Toxins, Biological / Q000378:metabolism*
D047908:Intracellular Signaling Peptides and Proteins; D008387:Marine Toxins; D052998:Microcystins; D010456:Peptides, Cyclic; D011994:Recombinant Proteins; D014118:Toxins, Biological; C063998:nodularin; D019319:Okadaic Acid; D019703:Calcineurin; C509249:PTPN6 protein, human; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C509250:Ptpn6 protein, rat; C057862:cyanoginosin LR