Improving Automated Protein Engineering Workflows with State-of-the-Art Plate Reading Capability
Abstract
Controlling the selectivity of chemical reactions and conducting these reactions in complex aqueous media, including unprocessed environmental or medical samples, could greatly expand our ability to analyze and synthesize chemicals in a non-laboratory setting. Metalloenzymes and artificial metalloenzymes (ArMs), the latter created by linking synthetic metal catalysts to protein scaffolds, could achieve these goals by enabling transition metal catalysis under conditions that would lead to inhibition or decomposition of free metal complexes. Moreover, protein scaffolds can be engineered to temporally control the reactivity of enzymes so that they are only active upon application of a stimulus. The Army Research Office has funded my groupÕs efforts in both of these areas through individual and MURI awards (W911NF1410334, W911NF1910074, W911NF1810200). Extensive protein engineering and directed evolution is required to optimize enzymes and ArMs for all of these applications. This process is greatly accelerated by laboratory automation, and previous DURIP funding was used to procure such a system for my laboratory (W911NF1510334). An integral part of this system is the plate readerÑthe analytical center of the system that is responsible for a broad range of high throughput assays to evaluate improved protein function. As the protein engineering required for the ARO-funded research noted above has advanced, the absorption and fluorescence spectroscopy capabilities of our ten-year-old plate reader are no longer adequate. This proposal describes the integration into our automation platform of a modern plate reader capable of making absorption, fluorescence, and luminescence measurements with greatly increased speed and sensitivity, compatibility with custom optics, and other essential features. This instrument will uniquely enable a broad range of fully-automated assays that are not possible using our existing plate reader and thereby accelerate our efforts to engineer enzymes and ArMs with improved catalytic activity and stimuli responsiveness.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Jan 27, 2022
- Source ID
- W911NF2210022
Entities
People
- Jared C Lewis
Organizations
- Army Contracting Command
- Indiana University
- United States Army