Subunit disassembly and inhibition of TNFα by a semi-synthetic bicyclic peptide.
Bottom Line: Macrocyclic peptides are potentially a source of powerful drugs, but their de novo discovery remains challenging.A 2.9 Å crystal structure of the M21/hTNFα complex reveals the peptide bound to a hTNFα dimer at a normally buried epitope in the trimer interface overlapping the binding site of a previously discovered small molecule ligand (SPD304), which also induces TNF trimer dissociation and synergizes with M21 in the inhibition of TNFα cytotoxicity.The discovery of M21 underlines the potential of semi-synthetic bicyclic peptides as ligands for the discovery of cryptic epitopes, some of which are poorly accessible to antibodies.
Affiliation: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK.Show MeSH
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Mentions: In order to investigate the nature of the time-dependent effect on TNFα inhibition by M21, we analyzed M21h peptide-TNFα complexes using liquid chromatography electrospray ionization time-of-flight mass spectrometry (LCT ESI-TOF MS), analytical ultracentrifugation (AUC) and size-exclusion chromatography multi-angle static light-scattering (SEC-MALS). MS spectra of TNFα showed a clear peak at around 52,508 Da corresponding to the trimeric form and stable over a long timeframe (4 weeks at 4°C). However, after incubation with bicyclic peptide M21h two peaks of 36,463 and 37,145 Da form, corresponding to a TNFα dimer with and without bound bicyclic peptide (Fig. 3A). After longer pre-incubation, TNFα trimers are converted quantitatively into dimers and monomers by M21h (Fig. 3B).Fig. 3.
Affiliation: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK.