Ion-Mediated Effects of Glycosylation on the Disordered Mucin Domain: Insights from Coarse-Grained Simulations.

Mucins are essential glycoproteins that form the backbone of mucus, a hydrogel protecting epithelial surfaces throughout the body. Their biophysical properties are governed by the densely glycosylated and highly disordered proline-threonine-serine (PTS) mucin domain, which becomes negatively charged by the addition of terminal sialic acid and sulfate groups to its glycans. The properties of mucins are further modulated by their interactions with cations, particularly sodium and calcium, which influence mucus expansion and viscoelasticity. Alterations in mucin glycosylation patterns or cation interactions contribute to the development of various pathological conditions. Modulating mucin's functional relationships to ameliorate these conditions requires first obtaining a detailed understanding of its structure, however, the large size, extensive disorder, and polymeric nature of mucins present significant challenges to their structural characterization. Here, we employed a coarse-grained modeling approach to investigate the effects of glycosylation, glycan charge, and salt concentration on mucin PTS organization. Using two different glycan structures, we explored how their interactions with monovalent (Na+) and divalent (Ca2+) cations affect the structural organization and ionic interactions of the PTS region. Our findings revealed that the presence of cations promoted tighter mucin conformations, with divalent cations inducing greater structural compaction than monovalent cations, consistently with their greater charge-shielding efficacy. Higher glycosylation levels and greater glycan charge densities enhance local glycan clustering and overall PTS structural expansion through their effects on cation binding and electrostatic repulsion. Furthermore, we examined the diffusion of charged peptides within mucin, demonstrating that peptide net charge affects both penetration and mobility. This study provides a comprehensive understanding of the structural organization and ionic interactions of mucin PTS, offering valuable insights into the molecular basis of mucin's protective properties and its role in health and disease.