Carla Zannella, Feliciana Real Fernandez, Federica Santoro, Rosa Bellavita, Bruno Casciaro, Paolo Rovero, Lucia Nencioni, Marta De Angelis, Anna De Filippis, Massimiliano Galdiero, Diego Brancaccio, Francesco Merlino, Paolo Grieco, Maria Luisa Mangoni, Alfonso Carotenuto
The study investigates temporin-derived antimicrobial peptides as broad-spectrum antiviral candidates against SARS-CoV-2. Surface plasmon resonance screening shows that temporin G ( TG ), temporin L ( TL ), and the nonhemolytic analog Pro 3 -TL bind the trimeric Spike protein with high affinity, following a two-state binding model. NMR confirms TG –Spike interactions and identifies Phe 2 as critical for binding. Docking simulations place the interaction at the NTD–RBD interface and reveal a cation−π interaction between TG Phe 2 and Spike Arg 357 . These insights informed the design of the chimeric analog RB-142 and four derivatives ( RB-143 to RB-146 ) incorporating bulkier aromatic residues. All analogs exhibit submicromolar binding affinities. Biological assays show low cytotoxicity and potent virucidal activity, with RB-146 demonstrating the strongest effect and a high therapeutic index. Mechanistic analyses indicate that RB-146 disrupts viral attachment and damages the viral envelope. The findings position RB-146 as a promising dual-mechanism antiviral candidate.