Amplifying the fluorescence of bilirubin enables the real-time detection of heme oxygenase activity |
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| Authors: | Roman Klemz Mir-Farzin Mashreghi Claudia Spies Hans-Dieter Volk Katja Kotsch |
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| Affiliation: | 1. Department of Anesthesiology and Intensive Care Medicine, Charité-Universitätsmedizin Berlin, Campus Virchow, 13353 Berlin, Germany;2. German Rheumatism Research Center, Berlin, Germany;3. Institute of Medical Immunology, Charité-Universitätsmedizin Berlin, Campus Mitte, 10117 Berlin, Germany;1. Laboratory of Mathematics and Physics, Ecole Centrale de Pékin, Beijing University of Aeronautics and Astronautics, Beijing 100191, China;2. The State Key Laboratory of Nonlinear Mechanics, Institute of Mechanics, Chinese Academy of Sciences, Beijing 100190, China;3. School of Energy and Power Engineering, Beijing University of Aeronautics and Astronautics, Beijing 100191, China;1. Department of Urology, Mayo Clinic, Rochester, MN;2. Department of Oncology, Mayo Clinic, Rochester, MN;1. Inserm U995, University of Lille Nord de France, Lille, France;2. Inserm ATIP-AVENIR, Denis Diderot University, Paris, France;3. Hepatology Unit, Claude Huriez Hospital, CHRU Lille, Lille, France;4. Intercommunal Hospital, Créteil, France;5. Inserm U866/Digestive Cancer Registry, Faculty of Medicine, Dijon, France;6. University Hospital, Burgundy University, Dijon, France;7. Hepato-Gastroenterology Unit, Jolimont Hospital, Haine-Saint-Paul, Belgium;8. Digestive Surgery and Liver Transplantation Unit, Claude Huriez Hospital, CHRU Lille, Lille, France;2. Department of Radiotherapy Physics, University College London Hospital, London, UK |
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| Abstract: | Heme oxygenases (HO) are the rate-limiting enzymes in the degradation of heme to equimolar amounts of antioxidant bile pigments, the signaling molecule carbon monoxide, and ferric iron. The inducible form HO-1 confers protection on cells and tissues that mediates beneficial effects in many diseases. Consequently, measurement of the enzymatic activity is vital in the investigation of the regulatory role of HO. Here we report that the fluorescence characteristics of bilirubin in complex with serum albumin can be used for the real-time detection of HO activity in enzymatic kinetics measurements. We characterized the enzymatic activity of a truncated human HO-1 and measured the HO activity for various cell types and organs, in either the basal naive or the HO-1-induced state. The bilirubin-dependent increase in fluorescence over time monitored by this assay facilitates a very fast, sensitive, and reliable measurement of HO activity. Our approach offers the basis for a highly sensitive high-throughput screening, which provides, inter alia, the opportunity to discover new therapeutic HO-1-inducing agents. |
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