This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are modeling the binding of endopolygalacturonase II from Aspergillus niger to its polygalacturonic acid (pectin) from cell walls using Amber and Glycam forcefields. In this investigation, there are important amino acid and carbohydrate residues that play a key role in binding, conformational change and catalysis. Employing post-simulational analysis with the MMPBSA approach, we are evaluating substrate binding free energies in solution. Employing computational alanine-scanning, will allow us to mutate various residues and observe their importance due to their absence based on charge, bulkiness and hydrophobicity. Future objectives are to correlate the solvent accessible surface found computationally with mass spectrometry and deuterium exchange. Deuterium exchange will give us insight on important residue interactions as well as a solvent accessible surface in the presence and absence of the polygalacturonic acid substrate. Other objectives include investigating the binding of the pectin substrate to enzyme in the presence of the PGIP inhibitor and the effects of the inhibitor on the enzyme/substrate complex.
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