Dynamics implications of glycan association with ferritin for nanocarrier design

Ferritin, a naturally occurring iron-storage protein, has emerged as a promising drug delivery platform. Previous studies have mainly focused on loading antigens onto the surface or inside the ferritin cage; however, investigations targeting the ferritin itself and its modification remain limited. Here, we investigated the atomistic effects of glycan association with ferritin monomers. Glycan association with ferritin was investigated as a surface-decoration approach, as it can improve functionality while preserving the native structural properties of the ferritin nanocage. Extensive molecular modelling and MD simulations were carried out to investigate four distinct glycan/glycan-derived residues separately on a ferritin monomer. N- and O-linked glycosylated ferritin were also modeled to distinguish non-covalent glycan association from covalent glycosidic modification. Docking results showed that the two glycan residues, Fmoc-L-Thr(α-D-Man(Ac) 4 )–OH and Man1-α-6[Man1-α-3]Man-β-O-pNp, represent higher binding affinity to ferritin. Dynamics of the glycated systems showed that GDP-mannose-bound ferritin displayed lower RMSD values compared to the other glycan-derived residues and is therefore the least flexible. It is followed by α-D-Man1-6α-D-Man1-benzyl and Fmoc-L-Thr(α-D-Man(Ac) 4 )–OH, while Man1-α-6[Man1-α-3]Man-β-O-pNp showed higher flexibility compared to the other glycan-derived residues. Overall, the glycan-derived residues with a smaller size and fewer units show reduced flexibility. RMSF results indicated that these glycan/glycan-derived residues induce localized changes in the flexibility of the outer surface of ferritin but do not affect the stability of the ferritin core or its functional regions. PCA and DCCM analyses further demonstrated that glycan residue binding does not lead to significant alterations in the dynamic behavior of ferritin, and the overall dynamics remain stable. The modeled glycan/glycan-derived residues were mainly localized to solvent-exposed regions of the ferritin monomer. The N- and O-linked glycosylated systems exhibited increased flexibility compared with all glyco-modified systems, supporting the idea that in the specific models examined here, the multi-site O-linked glycosylation model exhibited greater flexibility than the glycan-associated models. These findings suggest that ferritin cage modification can serve as a promising strategy for enhancing the functionality of the native ferritin system.