Bioluminescence is a popular technique for whole animal imaging. This technology relies on enzymes (luciferases) that catalyze the oxidation of small molecule substrates (luciferins), releasing photons of light in the process. While several luciferases and luciferins have been identified in nature, relatively few have been optimized for routine use in biomedical research. We are aiming to expand the number of bioluminescent probes for imaging <i>in vivo </i>by crafting new luciferase-luciferin pairs. Toward this end, we synthesized luciferin analogs comprising pyridone motifs. These analogs emitted different colors of light when incubated with firefly luciferase, suggesting that they are useful for multi-component imaging. We also used directed evolution to identify mutant luciferases that could more efficiently process the pyridone luciferins. Several of the mutant luciferases outperform native luciferases in terms of photon output. The mechanisms for improved activity were also investigated. Collectively, this work provides both new imaging tools and insights into evolving next-generation bioluminescent probes.<br/>

Evolved luciferases improve activity with pyridone luciferins

253rd National Meeting (2017)

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