Crystal structures and mutagenesis of PPP-family ser/thr protein phosphatases elucidate the selectivity of cantharidin and novel norcantharidin-based inhibitors of PP5C
- Univ. of Alabama, Birmingham, AL (United States)
- Univ. of South Alabama, Mobile, AL (United States)
- Geisel School of Medicine at Dartmouth, Lebanon, NH (United States)
- Univ. of Kansas Specialized Chemistry Center, Lawrence, KS (United States)
- Lunenfeld-Tanenbaum Research Inst. at Mount Sinai Hospital, Toronto, ON (Canada); Univ. of Toronto, ON (Canada)
- Cornell Univ., Lemont, IL (United States). Northeastern Collaborative Access Team (NE-CAT)
Cantharidin is a natural toxin and an active constituent in a traditional Chinese medicine used to treat tumors. Cantharidin acts as a semi-selective inhibitor of PPP-family ser/thr protein phosphatases. Despite sharing a common catalytic mechanism and marked structural similarity with PP1C, PP2AC and PP5C, human PP4C was found to be insensitive to the inhibitory activity of cantharidin. To explore the molecular basis for this selectivity, in this work we synthesized and tested novel C5/C6-derivatives designed from quantum-based modeling of the interactions revealed in the co-crystal structures of PP5C in complex with cantharidin. Structure–activity relationship studies and analysis of high-resolution (1.25 Å) PP5C-inhibitor co-crystal structures reveal close contacts between the inhibitor bridgehead oxygen and both a catalytic metal ion and a non-catalytic phenylalanine residue, the latter of which is substituted by tryptophan in PP4C. Quantum chemistry calculations predicted that steric clashes with the bulkier tryptophan side chain in PP4C would force all cantharidin-based inhibitors into an unfavorable binding mode, disrupting the strong coordination of active site metal ions observed in the PP5C co-crystal structures, thereby rendering PP4C insensitive to the inhibitors. This prediction was confirmed by inhibition studies employing native human PP4C. Mutation of PP5C (F446W) and PP1C (F257W), to mimic the PP4C active site, resulted in markedly suppressed sensitivity to cantharidin. These observations provide insight into the structural basis for the natural selectivity of cantharidin and provide an avenue for PP4C deselection. The novel crystal structures also provide insight into interactions that provide increased selectivity of the C5/C6 modifications for PP5C versus other PPP-family phosphatases.
- Research Organization:
- Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Organization:
- Abraham Mitchell Cancer Research Fund; Mayer Mitchell Annual Award for Excellence in Cancer Research; National Institutes of Health (NIH); Molecular Libraries Probe Production Centers Network (MLPCN); American Cancer Society; Canadian Cancer Society Research Institute (CCSRI); USDOE Office of Science (SC)
- Grant/Contract Number:
- U54HG005031; IRG-82-003-30; RO3MH05702; R21NS071553; AC02-06CH11357
- OSTI ID:
- 1251235
- Alternate ID(s):
- OSTI ID: 1425684
- Journal Information:
- Biochemical Pharmacology, Vol. 109, Issue C; ISSN 0006-2952
- Publisher:
- ElsevierCopyright Statement
- Country of Publication:
- United States
- Language:
- ENGLISH
Web of Science
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journal | December 2018 |
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journal | September 2016 |
Serine/threonine phosphatase 5 (PP5C/PPP5C) regulates the ISOC channel through a PP5C-FKBP51 axis
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journal | December 2017 |
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