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Antivitamin B12 Inhibition of the Human B12-Processing Enzyme CblC: Crystal Structure of an Inactive Ternary Complex with Glutathione as the Cosubstrate |
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| Authors: | Dr. Markus Ruetz Aranganathan Shanmuganathan Dr. Carmen Gherasim Agnes Karasik Dr. Robert Salchner M. Sc. Christoph Kieninger Dr. Klaus Wurst Prof. Ruma Banerjee Prof. Markos Koutmos Prof. Bernhard Kräutler |
| Affiliation: | 1. Institute of Organic Chemistry and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria University of Michigan Medical School, Ann Arbor, USA;2. Department of Biochemistry, Uniformed Services University of the Health Sciences, Bethesda, USA;3. University of Michigan Medical School, Ann Arbor, USA Current address: Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA;4. Institute of Organic Chemistry and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria Current address: Watercryst GmbH & Co, Kematen, Austria;5. Institute of Organic Chemistry and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria;6. Institute of General, Inorganic and Theoretical Chemistry, University of Innsbruck, Austria;7. University of Michigan Medical School, Ann Arbor, USA |
| Abstract: | B12 antivitamins are important and robust tools for investigating the biological roles of vitamin B12. Here, the potential antivitamin B12 2,4-difluorophenylethynylcobalamin (F2PhEtyCbl) was prepared, and its 3D structure was studied in solution and in the crystal. Chemically inert F2PhEtyCbl resisted thermolysis of its Co−C bond at 100 °C, was stable in bright daylight, and also remained intact upon prolonged storage in aqueous solution at room temperature. It binds to the human B12-processing enzyme CblC with high affinity (KD=130 nm ) in the presence of the cosubstrate glutathione (GSH). F2PhEtyCbl withstood tailoring by CblC, and it also stabilized the ternary complex with GSH. The crystal structure of this inactivated assembly provides first insight into the binding interactions between an antivitamin B12 and CblC, as well as into the organization of GSH and a base-off cobalamin in the active site of this enzyme. |
| Keywords: | antivitamins enzyme inhibitors glutathione protein crystallography vitamin B12 |