Revealing two distinct molecular binding modes in polyethyleneimine-DNA polyplexes using infrared spectroscopy.

In this study, we use infrared spectroscopy to investigate the molecular binding modes of DNA with linear and branched polyethylenimine (LPEI and BPEI). PEI-based polymers are widely studied as non-viral gene delivery vectors, but their low transfection efficiency limits their clinical success. One key factor affecting their performance is how they bind DNA as it directly impacts the packaging, protection, and release of the cargo in cells. While PEI-DNA binding has traditionally been viewed through the lens of electrostatics, computational models suggest additional binding mechanisms may be involved. Our findings reveal that LPEI and BPEI exhibit two distinct molecular binding modes, which influence DNA packaging into polyplexes. Identifying these binding modes provides critical insights into polymer complexation mechanisms to nucleic acids that can guide the rational design of more efficient and versatile PEI-based gene delivery systems.