A novel tag-free probe for targeting molecules interacting with a flavonoid catabolite |
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Authors: | Sayaka Nakashima Zhe Liu Yuya Yamaguchi Shunya Saiki Shintaro Munemasa Toshiyuki Nakamura Yoshiyuki Murata Yoshimasa Nakamura |
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Affiliation: | Graduate School of Environmental and Life Science, Okayama University, Okayama 700-8530, Japan |
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Abstract: | 3,4-Dihydroxyphenylacetic acid (DOPAC) is one of the colonic microflora-produced catabolites of quercetin 4′-glucoside (Q4′G). Although the interaction of DOPAC with cellular proteins might be involved in its biological activity, the actual proteins have not yet been identified. In this study, we developed a novel tag-free DOPAC probe to label the targeted proteins by the copper(I)-catalyzed azide alkyne cycloaddition (CuAAC) and verified its efficacy. Various labeled proteins were detected by the DOPAC probe with the azide labeled biotin and a horseradish peroxidase (HRP)-streptavidin complex. Furthermore, a pulldown assay identified Keap1 and aryl hydrocarbon receptor (AhR) as the target proteins for the phase 2 enzyme up-regulation. |
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Keywords: | Quercetin 3,4-dihydroxyphenylacetic acid (DOPAC) Click chemistry Tag free probe Keap1 |
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