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Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules
Published on: July 25, 2013
Phosphoester hydrolysis using a de novo designed protein: A combined experimental and computational study
Dona Imanga Upamadi Edirisinghe1, Prerana Dash2, Jiawen Yang3
1Department of Chemistry, Syracuse University, 111 College Place, Syracuse, NY 13244, United States of America.
None:
Metallohydrolases use metal cations in their active sites to catalyze the hydrolytic cleavage of biological substrates. We used a simple model protein Due Ferri single chain (DFsc) in combination with Mn-Mn and Zn-Zn to study the effect of the nature of metal ions on its reactivity. The ICP-OES analysis confirmed the metal ion binding with desired ratios. The Mn2-DFsc showed the highest hydrolytic activity with a catalytic efficiency (kcat/Km) of 6.5 M-1 min-1. EPR analysis of the di‑manganese DFsc indicates that two manganese centers are antiferromagnetically coupled in a S = 0 ground state. Based on both experimental and computational results, a mechanism of phosphoester hydrolysis is proposed, in which the substrate binds to Mn terminally, and follows an associative/dissociative mechanism with hydrolysis assisted by nucleophilic attack of the bridging OH nucleophile. The findings from this study can be used to further optimize the DFsc scaffold for designing artificial enzymes for bioremediation of multiple hazardous molecules.
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