Molecular Modeling of Polyelectrolyte Structure, Complexation and Aggregation in Gene Delivery

• Main Author: Gallops, Caleb Edward
• Format: Dissertation
• Language: English
• Published: ProQuest Dissertations & Theses 01-01-2021
• Subjects: Acids; Aqueous solutions; Carbon; Chemistry; Efficiency; Gene therapy; Genetic engineering; Genetics; Hypotheses; Investigations; Lipids; Nitrogen; Physical chemistry; Physical properties; Physiology; Polyamines; Polymerization; Polymers; Quantum dots; Simulation; Vectors (Biology)
• ISBN: 9798535510408

Polyelectrolyte complexes formed from nucleic acids and synthetic polycations have been studied for their use in non-viral gene delivery. Polyethylenimine (PEI) is a highly-studied polycationic vector with a high transfection efficiency that has been linked with its pH-responsiveness. In this series of studies, we examine the structural characteristics of PEI in atomistic detail and investigate the formation of polyelectrolyte complexes/aggregates using coarse-grained molecular dynamics. Atomistic molecular dynamics simulations of a linear 40mer PEI chain were performed for 9 protonation states and various NaCl concentrations to examine how the structure of PEI depends on pH and salt concentration. PEI continuously expands as it transitions from being unprotonated to fully protonated; however, we observe that two different regimes underlie this expansion. Sparsely protonated chains behave as weakly charged polyelectrolytes whose expansion is associated with the reduction of intrachain hydrophobic interactions. In contrast, the expansion of densely protonated chains with increased protonation involves increasing chain stiffness and breaking intrachain hydrogen bonds. The weakly-to-highly charged transition occurred at ~40% protonation, suggesting it may occur in endosomal conditions. These results provide a microscopic picture of changes in PEI structure during the gene delivery process. Coarse-grained molecular dynamics simulations are performed to examine the impact of chain length and polyanion stiffness on polyplex formation and aggregation. Polyplexes containing single polyanion chain fall into three structural regimes depending on polyanion stiffness: flexible polyanions form collapsed complexes, semiflexible polyanions form various morphologies including toroids and hairpins, and stiff polyanions form rod-like structures. Polyplex size generally decreases as polycation length increases. Aggregation (i.e., formation of complexes containing multiple polyanions) is observed in some simulations containing multiple polyanions and an excess of short polycations. Aggregation is observed to only occur for semiflexible and stiff polyanions and is promoted by shorter polycation lengths. Simulations of short, stiff polyanions condensed by long polycations are used as a model for siRNA gene delivery complexes. These simulations show multiple polyanions are spaced out along the polycation with polyanion-polyanion interactions, usually limited to overlapping chain ends. These structures differ from aggregates of longer polyanions in which the polyanions are packed together in parallel, forming bundles.