Atomistic insights into the structure, dynamics, and regulation of integral membrane proteins

With the rapid advances in GPU computing, cryo-EM structural biology, and deep learning–based structure prediction, Molecular dynamics (MD) simulations have become an indispensable tool for uncovering atomistic mechanisms that govern biomolecular function. The unique ability of MD to resolve molecular motions at high spatial and temporal resolution offers mechanistic insights into dynamic interactions that are often inaccessible to static structural methods alone. Cell membranes and their associated proteins play essential roles in fundamental cellular processes and are implicated in numerous diseases. Understanding the dynamics and regulation of membrane proteins is therefore critical for both basic biology and drug discovery. In this dissertation, under the guidance of Professor Emad Tajkhorshid, MD simulations—integrated with enhanced sampling, structural modeling, and free energy calculations—were employed to investigate conformational dynamics, lipid modulation, and domain coupling mechanisms in membrane protein systems. In the first study, we present a complete structural model of the SARS-CoV-2 spike protein, which forms the basis for the high pathogenicity and transmissibility of the virus. The full-length spike structure is constructed by integrating multiple advanced modeling techniques, incorporating large unresolved regions and experimentally characterized post-translational modifications. With microsecond-scale MD simulations, we show how glycan–glycan and glycan–lipid interactions broaden the protein’s dynamical range and thereby, its effective interaction with the surface receptors on the host cell, highlighting functional roles of glycans beyond surface shielding. My second project focuses on the transport mechanism of the human anion exchanger 1 (AE1), a membrane transporter that mediates rapid exchange of bicarbonate and chloride ions across the red blood cell membrane. We first identified the crucial residues involved in the substrate binding in both outward-facing (OF) and inward-facing (IF) states. Then, through the use of enhanced sampling techniques combining steered MD, string method and umbrella sampling, which can accelerate the sampling of rare events in the simulation, we explored the conformational transition pathway of AE1 between OF and IF states, which follows the alternating access mechanism, and characterized the free energy profiles along the path. Free energy profiling revealed the role of specific lipids in lowering the transition barrier, a finding supported by experimental validation, emphasizing the regulatory effect of lipid–protein interactions in AE1’s transport cycle. The human α7 nicotinic acetylcholine receptor (nAChR) is a key ligand-gated ion channel involved in cholinergic signaling throughout the nervous system. Its function is closely associated with the binding of neurotransmitters to its extracellular domain, which triggers conformational changes that open the distant transmembrane pore, allowing the influx of cations and leading to changes in membrane potential. In the final section, we investigated the allosteric domain coupling mechanism in the α7 nAChR which links the ligand binding to pore opening. Using structural modeling of mutants with insertions, complemented by ligand-binding assays and validated with long-timescale MD simulations, we revealed that single-residue insertions in the pre-M1 linker increase inter-domain spacing and disrupt the domain arrangement. Free energy calculations and network analysis further confirmed that these structural perturbations destabilize the native coupling geometry, weakening the dynamic correlation between the extracellular and transmembrane domains. Together, this thesis demonstrates the power of MD simulations to complement and extend experimental observations, revealing the structural, thermodynamic, and dynamic determinants of membrane protein function. These findings provide a computational framework for deeper mechanistic understanding of membrane proteins and their modulation in physiological and pathological contexts.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biophysics & Quant Biology

Grantor

University of Illinois Urbana-Champaign

Year dc:date

2025

Author dc:creator

• Chen, Tianle

Contributors dc:contributor

• Tajkhorshid, Emad
• Grosman, Claudio
• Pogorelov, Taras
• Wu, Nicholas C

• Molecular dynamics (MD) simulations
• Membrane proteins
• Conformational dynamics
• Lipid–protein interactions
• Free energy calculations
• Enhanced sampling
• SARS-CoV-2 spike protein
• Anion exchanger 1 (AE1)
• Alternating access mechanism
• α7 nicotinic acetylcholine receptor (nAChR)
• Domain coupling.

Statement dc:rights

• Copyright 2025 Tianle Chen

Language dc:language

en

Handle dc:identifier

https://hdl.handle.net/2142/132734

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/132734