title: Kinetics and thiol requirements of iodothyronine 5'-deiodination are tissue-specific in common carp (Cyprinus carpio L.).
abstract: Iodothyronine deiodinases determine the biological activity of thyroid hormones. Despite the homology of the catalytic sites of mammalian and teleostean deiodinases, in-vitro requirements for the putative thiol co-substrate dithiothreitol (DTT) vary considerably between vertebrate species. To further our insights in the interactions between the deiodinase protein and its substrates: thyroid hormone and DTT, we measured enzymatic iodothyronine 5'-deiodination, Dio1 and Dio2 mRNA expression, and Dio1 affinity probe binding in liver and kidney preparations from a freshwater teleost, the common carp (Cyprinus carpio L.). Deiodination rates, using reverse T3 (rT3, 3,3',5'-triiodothyronine) as the substrate, were analysed as a function of the iodothyronine and DTT concentrations. In kidney rT3 5'-deiodinase activity measured at rT3 concentrations up to 10 μM and in the absence of DTT does not saturate appreciably. In the presence of 1mM DTT, renal rT3 deiodination rates are 20-fold lower. In contrast, rT3 5'-deiodination in liver is potently stimulated by 1mM DTT. The marked biochemical differences between 5'-deiodination in liver and kidney are not associated with the expression of either Dio1 or Dio2 mRNA since both organs express both deiodinase types. In liver and kidney, DTT stimulates the incorporation of N-bromoacetylated affinity labels in proteins with estimated molecular masses of 57 and 55, and 31 and 28 kDa, respectively. Although primary structures are highly homologous, the biochemistry of carp deiodinases differs markedly from their mammalian counterparts.
authors.affiliation: Department of Organismal Animal Physiology, Institute for Water and Wetland Research, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, Box 30, 6525 AJ Nijmegen, The Netherlands. p.klaren@science.ru.nl
authors.forename: Peter H M
authors.initials: PH
authors.lastname: Klaren
authors.forename: Edwin J W
authors.initials: EJ
authors.lastname: Geven
authors.forename: Anika
authors.initials: A
authors.lastname: Nagelkerke
authors.forename: Gert
authors.initials: G
authors.lastname: Flik
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002347:Carps / Q000235:genetics / Q000378:metabolism*; D004229:Dithiothreitol / Q000494:pharmacology; D020869:Gene Expression Profiling; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D007453:Iodide Peroxidase / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000187:drug effects / Q000201:enzymology; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D009928:Organ Specificity / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D014020:Tissue Extracts / Q000378:metabolism; D014285:Triiodothyronine, Reverse / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D013438:Sulfhydryl Compounds; D014020:Tissue Extracts; D014285:Triiodothyronine, Reverse; D007453:Iodide Peroxidase; D004229:Dithiothreitol