A multivalent TAT–arginine-biodynamer conjugate targeting the bacterial cell envelope via specific membrane interactions

Antimicrobial resistance is a global crisis driven by a scarce pipeline of new antibiotics. A major contributor is the intrinsic resistance conferred by the bacterial envelope, highlighting the need for innovative molecules for improved therapies. In this study, TAT-ArgBD, a conjugate of the cell-penetrating TAT peptide and arginine biodynamer (ArgBD), serves in vitro as a multivalent macromolecular antibiotic and synergist. TAT-ArgBD rapidly kills 99.9% of Pseudomonas aeruginosa at 32 µg/mL within 1 h, outperforming colistin, and shows minimum inhibitory concentrations (MICs) of 2-8 µg/mL against Acinetobacter baumannii and Staphylococcus aureus. Notably, it potentiates antibiotics such as novobiocin, chloramphenicol, and imipenem, leading to lowered MICs up to 256-fold. Notably, novobiocin, typically active only against Gram-positive bacteria, showed activity against Gram-negative bacteria when combined with TAT-ArgBD. Mechanistic studies suggest TAT-ArgBD antimicrobial and synergistic actions result from preferential binding to POPG and cardiolipin. This interaction induces bacterial membrane pore formation by adopting an α-helical conformation in the presence of bacterial lipids. With a favorable in vitro safety profile, a membranolytic index > 64 and low mammalian cell toxicity at effective bactericidal concentrations, TAT-ArgBD's potential to enhance antibiotic efficacy, as well as function as a stand-alone treatment, supports further preclinical evaluation as an antimicrobial adjuvant.