Impact of Acetylation,\nSuccinylation, and pH on DNA\nPackaging in PEI-Based Polyplexes

Polyethylenimine (PEI) is a widely used cationic polymer\nfor nonviral\ngene delivery, often modified to enhance transfection efficiency and\nreduce cytotoxicity. This study investigates how acetylation, succinylation\n(acPEI and zPEI), and pH influence the internal DNA packaging of polyplexes.\nBoth modifications alter physicochemical properties, leading to complexes\nthat decondense more readily with increasing modification. X-ray scattering\nreveals that high acetylation produces loosely packed DNA, while succinylation\nunexpectedly tightens DNA packing at higher modification levels. Polyplexes\nformed at low pH (pH 4) are more stable and tightly packed than those\nformed at pH 7.5. Acidifying polyplexes initially formed at pH 7.5\ninduces structural rearrangement to tighter DNA packing accompanied\nby significant PEI release, providing direct evidence for models where\nfree PEI aids endosomal escape. These findings challenge conventional\nassumptions about PEI behavior and offer new insights into DNA packaging,\nemphasizing tailored polymer modifications and pH conditions to optimize\ngene delivery.