title: Identification of iopanoic acid as substrate of type 1 deiodinase by a novel nonradioactive iodide-release assay.
abstract: Enzymatic 5'- and 5-deiodination are key reactions for local and systemic activation and inactivation of iodothyronines and thyronamines. Expression of the three deiodinase (DIO) isoenzymes is regulated by a number of parameters, including thyroid status, genotype, micronutrient availability, and disease-related signaling. In addition, DIO are potential targets of pharmacological as well as environmentally derived substances, which might affect their enzymatic activity (endocrine disruptors). With the classical DIO activity assay, testing depends on the availability of radioactively labeled substrates (e.g. (125)I-rT(3)) to monitor the release of radioactive iodide. Recently, liquid chromatography-tandem mass spectrometry was described as an alternative method apparently resolving this limitation. However, it has a high demand in technical equipment and analytical routine and is limited in sample number by considerable measuring time. We therefore combined the classical deiodination assay with an easily accessible photometric method taking advantage of the Sandell-Kolthoff reaction for measuring iodide release. In brief, iodine works as a catalyst within this redox reaction between Ce(4+) and As(3+) leading to an acceleration of destaining. Furthermore, the protocol was adapted to minimize handling effort and time consumption. Because this method is not dependent on radioactivity, it expands the substrate spectrum of the classical method. Suitability of this assay was tested with tissue samples from animal experiments (hepatic Dio1 activity in hypo- and hyperthyroid mice) and established DIO inhibitors. As a new but not unexpected finding, the alleged inhibitor iopanoic acid turned out to be a DIO substrate. This finding was confirmed by liquid chromatography-tandem mass spectrometry, and its potential clinical impact requires further studies.
authors.affiliation: Institut für Experimentelle Endokrinologie, Charité-Universitätsmedizin Berlin, Augustenburger Platz 1, D-13353 Berlin, Germany. kostja.renko@charite.de
authors.forename: Kostja
authors.initials: K
authors.lastname: Renko
authors.forename: Carolin S
authors.initials: CS
authors.lastname: Hoefig
authors.forename: Franziska
authors.initials: F
authors.lastname: Hiller
authors.forename: Lutz
authors.initials: L
authors.lastname: Schomburg
authors.forename: Josef
authors.initials: J
authors.lastname: Köhrle
mesh_terms: D000818:Animals; D007453:Iodide Peroxidase / Q000378:metabolism*; D007480:Iopanoic Acid / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D051379:Mice; D053719:Tandem Mass Spectrometry; D013961:Thyroid Gland / Q000378:metabolism*
chemical_list: C109812:iodothyronine deiodinase type I; D007453:Iodide Peroxidase; D007480:Iopanoic Acid