title: Functional analysis of iodotyrosine deiodinase from drosophila melanogaster.
abstract: The flavoprotein iodotyrosine deiodinase (IYD) was first discovered in mammals through its ability to salvage iodide from mono- and diiodotyrosine, the by-products of thyroid hormone synthesis. Genomic information indicates that invertebrates contain homologous enzymes although their iodide requirements are unknown. The catalytic domain of IYD from Drosophila melanogaster has now been cloned, expressed and characterized to determine the scope of its potential catalytic function as a model for organisms that are not associated with thyroid hormone production. Little discrimination between iodo-, bromo-, and chlorotyrosine was detected. Their affinity for IYD ranges from 0.46 to 0.62 μM (Kd) and their efficiency of dehalogenation ranges from 2.4 - 9 x 103M-1s-1(kcat/Km). These values fall within the variations described for IYDs from other organisms for which a physiological function has been confirmed. The relative contribution of three active site residues that coordinate to the amino acid substrates was subsequently determined by mutagenesis of IYD from Drosophila to refine future annotations of genomic and meta-genomic data for dehalogenation of halotyrosines. Substitution of the active site glutamate to glutamine was most detrimental to catalysis. Alternative substitution of an active site lysine to glutamine affected substrate affinity to the greatest extent but only moderately affected catalytic turnover. Substitution of phenylalanine for an active site tyrosine was least perturbing for binding and catalysis.
authors.affiliation: Department of Chemistry, Johns Hopkins University, 3400 N. Charles St, Baltimore, Maryland, 21218.
authors.forename: Abhishek
authors.initials: A
authors.lastname: Phatarphekar
authors.affiliation: Department of Chemistry, Johns Hopkins University, 3400 N. Charles St, Baltimore, Maryland, 21218.
authors.forename: Steven E
authors.identifier: 0000-0002-2292-2917
authors.initials: SE
authors.lastname: Rokita
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D020134:Catalytic Domain; D029721:Drosophila Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster; D007453:Iodide Peroxidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D013379:Substrate Specificity; D014443:Tyrosine / Q000031:analogs & derivatives / Q000737:chemistry
chemical_list: D029721:Drosophila Proteins; D014443:Tyrosine; D007453:Iodide Peroxidase
keywords: Drosophila; dehalogenase; deiodinase; flavoprotein; halotyrosine; iodotyrosine