(316a) Single-Molecule Characterization of Cazyme Protein Modules Adsorption to Multivalent Glucan Polymers like Cellulose | AIChE

(316a) Single-Molecule Characterization of Cazyme Protein Modules Adsorption to Multivalent Glucan Polymers like Cellulose


Chundawat, S. - Presenter, Rutgers University
Nemmaru, B., Rutgers University
Gnanakaran, G., Los Alamos National Laboratory
Lopez, C., Los Alamos National Laboratory
Lang, M., Vanderbilt University
Hilton, M., Vanderbilt University
Hackl, M., Rutgers, State University of New Jersey
Protein adsorption to multivalent carbohydrate (or glycan) derived ligands plays a critical, but poorly understood, role in both fundamental cellular (e.g., glycocalyx and viral protein binding) and industrial processes (e.g., cellulosic sugar-based biofuels production). However, we currently have a poor understanding of the molecular mechanisms driving protein adsorption to glucan polymers like cellulose. We also lack suitable experimental tools and predictive adsorption models that can suitably explain the heterogeneous and varying affinity multivalent protein-carbohydrate binding interactions. Here, we study the adsorption-desorption of a family 1 carbohydrate-binding module (CBM), and its conserved aromatic amino acid mutants, to model glucan polymers like cellulose using a combination of bulk ensemble and single-molecule protein-ligand binding methods. Our single-molecule force spectroscopy method allows us to directly probe the structure-function relationships driving CBM adsorption to glucan polymers. Analysis of the CBM-cellulose unbinding rupture forces and total bond lifetimes dataset has provided a molecular basis for distinct binding orientations for CBMs on ordered glucan polymer surfaces like cellulose and the corresponding binding affinity for each orientation. Molecular dynamics simulations provide further insights into the possibility of multiple binding orientations for CBM on cellulose surfaces, which is an often-neglected feature of these interactions. We also report ongoing work with the aromatic amino acid mutants that are critical to stabilizing specific binding orientations and how these binding orientations could vary for various Type-A and Type-B CBMs on cellulose allomorphs. Our findings allow us to generate a comprehensive picture of non-productive binding interaction mechanisms of CBMs with glucan polymers that will ultimately lead to the development of more efficient enzymes for cellulosic biomass deconstruction into fermentable sugars.