Structural and functional role of methylene linkers in self-coacervation-mediated membrane interaction and antiviral defense

Designing small-molecule-based (MW ≤ 1500) self-coacervates and tuning their dynamic properties holds great significance in the regulation of biological events. Herein, we designed a series of pKa tunable polyionic lipidated flavonoids (LFs), where the aromatic core of flavonoid and the acyl chains serve as stickers, the hydrophilic polyanionic moieties interact via multivalent COO‾…HOOC hydrogen bond formation, leading to coacervation at low concentrations (10 µM). Inspired by nature’s selection of amino acids (glutamic acid vs 2-amino hexane dioic acid) with the profound role of methylene units, we screened a library of LFs to explore the influence of methylene linker length (n = 0-3). FRAP studies suggest that shorter linkers (n = 0, 1) produce gel-like coacervates, whereas longer linkers (n = 2, 3) yield more fluidic droplets. The self-coacervates exhibited strong membrane association, resulting in the formation of 2D domains and/or nano-coacervate-like domains within the lipid bilayer. The domains MFEA (n = 2) are membrane-active, inhibit membrane fusion, and protect the cells from SARS-CoV-2 and influenza infections, highlighting the potential as a broad-spectrum antiviral. Our design of amphiphilic self-coacervates offers a versatile new platform for engineering functional materials and highlights the role of methylene linker length for tuning coacervate mechanics. Small-molecule-based self-coacervates exhibit tunable dynamic properties; however, liquid-liquid phase separation in such systems at low microcomolar concentrations remains underexplored. Here, the authors report the design and dynamic properties of flavonoid-based amphiphilic self-coacervates, the modulation of the critical coacervate concentration and membrane interactions to form domains — which order the membrane interface, inhibit membrane fusion, and could combat viral infections caused by enveloped viruses.