(527c) Single-Molecule Characterization of Protein Adsorption to Multivalent Glycan Polymers

Protein adsorption to multivalent carbohydrate (or glycan) derived ligands plays a critical, but poorly understood, role in both fundamental cellular (e.g., cell glycocalyx and viral protein binding) and industrial processes (e.g., cellulosic biofuel production). However, we currently have a poor understanding of the molecular mechanisms driving protein adsorption to complex glycan polymers. We also lack suitable experimental tools and predictive adsorption models that can explain the heterogeneous and high affinity multivalent protein-carbohydrate binding interactions. Here, we study the adsorption of native carbohydrate-binding proteins to model polysaccharides using complementary bulk and single-protein based adsorption techniques. Our single-protein adsorption method is an optical tweezers based bond lifetime assay following a modified methodology as first reported by Brady et al. This technique has allowed us to probe protein-glycan binding interactions at a range of rupture forces. Detailed analysis of the single-protein assay dataset revealed underlying trends that help explain the bulk ensemble observations. The range of bond lifetimes and forces reflect the underlying heterogeneity of available binding sites for multivalent carbohydrate-based ligands. These results provide an underlying molecular basis for using multi-site adsorption models to characterize protein adsorption to multivalent polysaccharides.

Brady, S. K., Sreelatha, S., Feng, Y., Chundawat, S. P. S. & Lang, M. J. Cellobiohydrolase 1 from Trichoderma reesei degrades cellulose in single cellobiose steps. Nature Communications. 6, 10149 (2015).