CHARMM γ-modified peptide nucleic acid parameterization and simulations for G-quadruplex invasion

Familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are neurodegenerative diseases genetically linked with a G₄C₂ hexanucleotide repeat expansion in the chromosome 9 open reading frame 72. The G-rich repeats are known to adopt G-quadruplex (GQ) tertiary structures under physiological conditions, co-existing in equilibrium with a hairpin (HP) structure. GQs have been suggested to sequester RNA binding proteins, causing toxic foci leading to RNA toxicity and potentially to the development of ALS/FTD. Gamma-modified peptide nucleic acids (γPNA) have the invasive properties of disrupting the GQ and HP tertiary structures while evading degradation in vivo, demonstrating the potential in becoming a therapeutic agent. Within our work, we model the complementary C₄G₂ γPNA modified with a diethylene glycol (DEG) on the γ-position. Parameters for the Chemistry at Harvard macromolecular mechanics (CHARMM) force field were developed using the CHARMM general force field (CGenFF) method for linkage between DEG and the γPNA backbone. Rotational scans were done on the linkage quantum mechanically (QM) by rotating the dihedral every 15° at the second-order Møller-plesset perturbation theory, MP2, using Pople’s 6-31+G* basis set. The rotation profile became our QM target data. Initial parameters derived from the CGenFF server for the linkage and were concatenated with existing parameters for the PNA backbone for molecular mechanics (MM) optimization. The initial force constant (K_χ) for the dihedral MM parameters were iteratively modified from 1.80 kcal/mol to 2.40 kcal/mol until convergence with the QM rotational scan. CHARMM-based molecular dynamics (MD) simulations were performed on the γPNA, HP, and GQ structures at the µs time scale to derive representative structures for docking identified by principal component analysis (PCA). We used HDOCK to dock representative structures of our γPNA to the HP and GQ representative structures for MD simulation. We further refined the γPNA/RNA complexes with 1 µs simulation, comparing the RMSF between our GQ without the γPNA and with the γPNA. Our MD simulations suggest that the complementary γPNA stabilizes the loops of the GQ with less fluctuation reported within the RMSF. Preliminary interpretations suggest stabilization of GQ may be the onset of invasion. Fundamentally, this work has implications in other neurodegenerative diseases and may potentially derive new therapeutics based on γPNAs.