Parallel Screening for Rapid Identification of Orthogonal Bioluminescent Tools.

Autor:	Rathbun CM; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Porterfield WB; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Jones KA; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Sagoe MJ; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Reyes MR; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Hua CT; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States., Prescher JA; Department of Chemistry, Department of Molecular Biology & Biochemistry, and Department of Pharmaceutical Sciences, University of California, Irvine Irvine, California 92697, United States.
Jazyk:	angličtina
Zdroj:	ACS central science [ACS Cent Sci] 2017 Dec 27; Vol. 3 (12), pp. 1254-1261. Date of Electronic Publication: 2017 Nov 15.
DOI:	10.1021/acscentsci.7b00394
Abstrakt:	Bioluminescence imaging with luciferase enzymes and luciferin small molecules is a well-established technique for tracking cells and other biological features in rodent models. Despite its popularity, bioluminescence has long been hindered by a lack of distinguishable probes. Here we present a method to rapidly identify new substrate-selective luciferases for multicomponent imaging. Our strategy relies on parallel screening of luciferin analogues with panels of mutant enzymes. The compiled data set is then analyzed in silico to uncover mutually orthogonal sets. Using this approach, we screened 159 mutant enzymes with 12 luciferins. Thousands of orthogonal pairs were revealed with sufficient selectivity for use in biological environments. Over 100 pairs were validated in vitro , and three were applied in cell and animal models. The parallel screening method is both generalizable and scalable and will streamline the search for larger collections of orthogonal probes. Competing Interests: The authors declare no competing financial interest.
Databáze:	MEDLINE
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