Enzymatic modification of tRNAs: MiaB is an iron-sulfur protein.
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.
Pierrel
Fabien
F
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.
Björk
Glenn R
GR
Fontecave
Marc
M
Atta
Mohamed
M
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
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