Luciferases are a valuable biotechnological used in cell imaging and medical diagnostics. Bioluminescent imaging with luciferase has many advantages over traditional fluorescent protein imaging, but the luciferase is not optimized for many biotechnological functions. While previous research has attempted to address these shortcomings, researchers have struggled to find beneficial mutations that do not lead to destabilization or reduced activity. Here, we use directed evolution to modify the active site chemistry of a luciferase from <i>Photinus pyralis</i>. Our library was designed to include natural variation of red-shifted homologs and previous literature mutations around the luciferin-binding pocket. We also designed a multi-property screen to identify enzymes with beneficial mutations imparting redshifted emissions and/or lower substrate binding coefficients.

Efforts towards engineered luciferases with optimized function

251st National Meeting (2016)

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