1. Roseltide rT7 is a disulfide-rich, anionic, and cell-penetrating peptide that inhibits proteasomal degradation.
- Author
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Kam A, Loo S, Fan JS, Sze SK, Yang D, and Tam JP
- Subjects
- A549 Cells, Anti-Inflammatory Agents pharmacology, Antimicrobial Cationic Peptides, Antineoplastic Agents, Phytogenic pharmacology, Cysteine chemistry, Disulfides, Endocytosis, Flow Cytometry, Humans, Magnetic Resonance Spectroscopy, Microscopy, Confocal, Molecular Conformation, Plant Lectins, Plant Proteins chemistry, Proteomics, Structure-Activity Relationship, Ubiquitin chemistry, Cell-Penetrating Peptides pharmacology, Hibiscus chemistry, Plant Extracts chemistry, Proteasome Endopeptidase Complex metabolism, Proteasome Inhibitors pharmacology
- Abstract
Disulfide-rich plant peptides with molecular masses of 2-6 kDa represent an expanding class of peptidyl-type natural products with diverse functions. They are structurally compact, hyperstable, and underexplored as cell-penetrating agents that inhibit intracellular functions. Here, we report the discovery of an anionic, 34-residue peptide, the disulfide-rich roseltide rT7 from Hibiscus sabdariffa (of the Malvaceae family) that penetrates cells and inhibits their proteasomal activities. Combined proteomics and NMR spectroscopy revealed that roseltide rT7 is a cystine-knotted, six-cysteine hevein-like cysteine-rich peptide. A pair-wise comparison indicated that roseltide rT7 is >100-fold more stable against protease degradation than its S -alkylated analog. Confocal microscopy studies and cell-based assays disclosed that after roseltide rT7 penetrates cells, it causes accumulation of ubiquitinated proteins, inhibits human 20S proteasomes, reduces tumor necrosis factor-induced IκBα degradation, and decreases expression levels of intercellular adhesion molecule-1. Structure-activity studies revealed that roseltide rT7 uses a canonical substrate-binding mechanism for proteasomal inhibition enabled by an IIML motif embedded in its proline-rich and exceptionally long intercysteine loop 4. Taken together, our results provide mechanistic insights into a novel disulfide-rich, anionic, and cell-penetrating peptide, representing a potential lead for further development as a proteasomal inhibitor in anti-cancer or anti-inflammatory therapies., (© 2019 Kam et al.)
- Published
- 2019
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