2.1.1. System Construction

Coordinates for all systems were constructed using the CHARMM software [69–71] v. c41b2 with the CHARMM36 (C36) biomolecular force field for carbohydrates [56,61–63]. Of note, it has been shown that MD simulations can reproduce ring puckers observed by NMR [49–51], with one study demonstrating the capacity of the CHARMM36 force field to reproduce NMR data for an iduronate derivative in the context of a heparin analogue [51]. The initial conformation for an MD simulation of non-sulfated chondroitin 20-mer was fully extended, with glycosidic linkage dihedrals φ = −83.75◦ and ψ = −156.25◦ in all GlcAβ1-3GalNAc linkages and φ = −63.75◦ and ψ = 118.75◦ in all GalNAcβ1-4GlcA linkages. These glycosidic linkage dihedral angle values were found to be the most energetically favorable in MD-simulated, non-sulfated chondroitin disaccharides [54]. All other internal coordinates were taken from the force-field files. In this conformation, the 20-mer had an end-to-end distance of 101.8 Å and was solvated in a cubic periodic unit cell with an edge length of 124.3 Å (~63,000 water molecules). The explicit solvent consisted of the TIP3P water model [72,73], neutralizing Na+ counterions, and 140 mM sodium chloride.
