Date of Award

Spring 2025

Project Type

Dissertation

Program or Major

Chemistry

Degree Name

Doctor of Philosophy

First Advisor

Harish Vashisth

Second Advisor

Charles Zercher

Third Advisor

Krisztina Varga

Abstract

Molecular transport across membranes is an important biophysical process. Designing molecules capable of crossing cell membranes and targeting specific organelle is becoming necessary in treating many diseases and disorders. In recent decades, cell-penetrating peptides have been studied and utilized to deliver a variety of cargoes. My dissertation work focuses on studying the molecular details of the peptide translocation process across a cell membrane. Specifically, I investigated the dynamics of mitochondrial-targeting peptides and their mechanisms of translocation across the mitomembrane. I conducted molecular dynamics (MD) simulations of model mitochondrial-targeting peptides with distinct physical and chemical properties such as sequence lengths, functional groups in sidechains, and the locations of the conjugated molecules. I began my work by performing equilibrium all-atom MD simulations of peptide and membrane systems. From these simulations, I determined that my model peptides had affinities toward the model mitomembrane, primarily driven by electrostatic attraction between the cationic residues and the membrane anionic headgroups. I then studied the peptide translocation process across the mitomembrane in all-atom systems using non-equilibrium MD simulations. These studies revealed a barrier-mediated direct translocation process for these peptides. Peptides with distinct physical structures adopted different metastable states and unique conformations during their transport across the membrane. In addition to studying the dynamics of peptide transport across the membrane via direct translocation in all-atom peptide and membrane systems, I also investigated peptide translocation under the influence of an external electric field to understand the role of membrane potential in peptide transport. Specifically, I conducted field-driven coarse-grained MD simulations on model peptides and membrane systems under various applied field strengths. In these field-driven MD simulations, peptides translocated via a pore formation mechanism which occurred independently of the presence of peptides. I also found that increasing the field strength decreases the translocation time, thereby suggesting the role of membrane potential in peptide translocation. These studies highlighted other mechanisms and pathways for peptide translocation across the mitomembrane. Overall, the work in my dissertation emphasized the importance of peptide design in facilitating interactions with and transport across the membrane. Additionally, my studies revealed mechanisms underlying these biophysical processes. Findings from my dissertation work provide an enhanced mechanistic understanding of peptide therapeutics targeting mitomembranes, which is essential for designing further optimized peptides.

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