Force transfer in the glycosylated human CLEC-2-podoplanin PLAG interface

Platelet C-type lectin-like receptor 2 (CLEC-2) binds an O-glycosylated podoplanin PLAG-region peptide through a side-face interface. The crystal structure defines the local glycan and peptide interactions, while their respective contributions to interfacial stability and mechanical load transmission require further mechanistic resolution. Here, atomistic simulations of the experimentally resolved CLEC-2-podoplanin glycopeptide complex were used to examine how these two ligand components contribute to interfacial mechanics. The sialylated Thr52 glycan formed a persistent recognition anchor, whereas the podoplanin Glu47/Asp48 segment together with the CLEC-2 Arg107/Arg152/Arg157 region constituted the principal load-transfer pathway during normal pulling. Consistently, deglycosylation produced smaller changes in equilibrium contacts and constant-force survival lower bounds than mutations of the acidic peptide residues or receptor arginines. These results indicate a functional separation within the interface, in which the Thr52 glycan primarily stabilizes local recognition, while interactions between the acidic peptide residues and CLEC-2 arginines dominate mechanical load transmission.

The 3WSR CLEC-2-podoplanin glycopeptide complex was simulated in GROMACS using CHARMM36m protein and CHARMM carbohydrate parameters. Independent equilibrium replicas were used to assess complex stability, periodic-image separation, and peptide and glycan contact occupancies. Constant-velocity SMD was retained as a relative forced separation comparison, while constant-force SMD was used as a lower-bound survival comparison under normal load. The analysis includes WT, deglycosylated, podoplanin E47A/D48A, CLEC-2 R152A/R157A, and single CLEC-2 R107A, R152A, and R157A systems.