title: Intracellular Generation of a Diterpene-Peptide Conjugate that Inhibits 14-3-3-Mediated Interactions.
abstract: Synthetic agents that disrupt intracellular protein-protein interactions (PPIs) are highly desirable for elucidating signaling networks and developing new therapeutics. However, designing cell-penetrating large molecules equipped with the many functional groups necessary for binding to large interfaces remains challenging. Here, we describe a rational strategy for the intracellular oxime ligation-mediated generation of an amphipathic bivalent inhibitor composed of a peptide and diterpene natural product, fusicoccin, which binds 14-3-3 protein with submicromolar affinity. Our results demonstrate that co-treatment of cells with small module molecules, the aldehyde-containing fusicoccin 1 and the aminooxy-containing peptide 2, generates the corresponding conjugate 3 in cells, resulting in significant cytotoxicity. In contrast, chemically synthesized 3 is not cytotoxic, likely due to its inability to penetrate cells. Compound 3, but not 1 or 2, disrupts endogenous 14-3-3/cRaf interactions, suggesting that cell death is caused by inhibition of 14-3-3 activity. These results suggest that intracellular generation of large-sized molecules may serve as a new approach for modulating PPIs.
authors.affiliation: Institute for Chemical Research and Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University , Gokasho, Uji, Kyoto 611-0011, Japan.
authors.forename: Prakash
authors.initials: P
authors.lastname: Parvatkar
authors.affiliation: Institute of Scientific and Industrial Research, Osaka University , 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan.
authors.forename: Nobuo
authors.initials: N
authors.lastname: Kato
authors.affiliation: Institute for Chemical Research and Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University , Gokasho, Uji, Kyoto 611-0011, Japan.
authors.forename: Motonari
authors.initials: M
authors.lastname: Uesugi
authors.affiliation: Institute for Chemical Research and Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University , Gokasho, Uji, Kyoto 611-0011, Japan.
authors.forename: Shin-Ichi
authors.initials: S
authors.lastname: Sato
authors.affiliation: Institute for Chemical Research and Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University , Gokasho, Uji, Kyoto 611-0011, Japan.
authors.forename: Junko
authors.initials: J
authors.lastname: Ohkanda
mesh_terms: D048948:14-3-3 Proteins / Q000737:chemistry*; D004224:Diterpenes / Q000737:chemistry*; D057809:HEK293 Cells; D006801:Humans; D062105:Molecular Docking Simulation; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding
chemical_list: D048948:14-3-3 Proteins; D004224:Diterpenes; D010455:Peptides