Submission Date

4-28-2025

Document Type

Paper- Restricted to Campus Access

Department

Biology

Adviser

Amanda Reig

Committee Member

Erica Gorenberg

Committee Member

Rebecca Roberts

Department Chair

Denise Finney

Project Description

G4DFsc is a de novo protein designed to mimic the metal-binding active sites of natural non-heme diiron enzymes. Through site-directed mutagenesis, specific amino acids can be targeted to affect conformational changes in the active site. In this study, H77D or H77E mutations were made in the E44H-G4DFsc plasmid to create an active site in which the histidine amino acids are trans to one another. Analysis of the mutated protein can further characterize the structure-function relationship of the protein’s hydrolase capabilities. The H77D and H77E mutagenesis was successful. Both proteins were produced and purified, which allowed for characterization of their biophysical properties. Future studies will include analysis of their DNA cleavage activity.

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