title: Enzymatic modification of tRNAs: MiaB is an iron-sulfur protein.
abstract: The product of the miaB gene, MiaB, from Escherichia coli participates in the methylthiolation of the adenosine 37 residue during modification of tRNAs that read codons beginning with uridine. A His-tagged version of MiaB has been overproduced and purified to homogeneity. Gel electrophoresis and size exclusion chromatography revealed that MiaB protein is a monomer. As isolated MiaB contains both iron and sulfide and an apoprotein form can chelate as much as 2.5-3 iron and 3-3.5 sulfur atoms per polypeptide chain. UV-visible and EPR spectroscopy of MiaB indicate the presence of a [4Fe-4S] cluster under reducing and anaerobic conditions, whereas [2Fe-2S] and [3Fe-4S] forms are generated under aerobic conditions. Preliminary site-directed mutagenesis studies suggest that Cys(157), Cys(161), and Cys(164) are involved in iron chelation and that the cluster is essential for activity. Together with the previously shown requirement of S-adenosylmethionine (AdoMet) for the methylthiolation reaction, the finding that MiaB is an iron-sulfur protein suggests that it belongs to a superfamily of enzymes that uses [Fe-S] centers and AdoMet to initiate radical catalysis. MiaB is the first and only tRNA modification enzyme known to contain an Fe-S cluster.
authors.affiliation: Laboratoire de Chimie et Biochimie des Centres Rédox Biologiques, Département de Biologie Moléculaire et Structurale-Chimie Biologie, UMR 5047 Commissariat à l'Energie Atomique (CEA)/CNRS/Université Joseph Fourier, Grenoble, France.
authors.forename: Fabien
authors.initials: F
authors.lastname: Pierrel
authors.forename: Glenn R
authors.initials: GR
authors.lastname: Björk
authors.forename: Marc
authors.initials: M
authors.lastname: Fontecave
authors.forename: Mohamed
authors.initials: M
authors.lastname: Atta
mesh_terms: D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003062:Codon / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D007501:Iron / Q000737:chemistry*; D008956:Models, Chemical; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012343:RNA, Transfer / Q000378:metabolism*; D013440:Sulfides / Q000737:chemistry*; D013466:Sulfurtransferases / Q000378:metabolism / Q000502:physiology*; D013997:Time Factors; D014466:Ultraviolet Rays; D014529:Uridine / Q000378:metabolism
chemical_list: D003062:Codon; D029968:Escherichia coli Proteins; D013440:Sulfides; D012343:RNA, Transfer; D007501:Iron; C499584:MiaB protein, E coli; D013466:Sulfurtransferases; C402430:isopentenyl-adenosine i6a thiotransferase; D003545:Cysteine; D014529:Uridine