TY - JOUR
T1 - Two Fe-S clusters catalyze sulfur insertion by radical-SAM methylthiotransferases
AU - Forouhar, Farhad
AU - Arragain, Simon
AU - Atta, Mohamed
AU - Gambarelli, Serge
AU - Mouesca, Jean Marie
AU - Hussain, Munif
AU - Xiao, Rong
AU - Kieffer-Jaquinod, Sylvie
AU - Seetharaman, Jayaraman
AU - Acton, Thomas B.
AU - Montelione, Gaetano T.
AU - Mulliez, Etienne
AU - Hunt, John F.
AU - Fontecave, Marc
N1 - Funding Information:
We thank R. Abramowitz and J. Schwanof for assistance with synchrotron data collection, B. Gibney for advice on Fe-S reconstitution for crystallization and R. Breslow for discussion of the reaction mechanism. We thank O. Hamelin for GC/MS analysis and J.-M. Moulis for providing 77Se (both from Chemistry and Biology of Metals Laboratory, Grenoble). This work was supported by the US National Institutes of Health Protein Structure Initiative grants U54-GM074958 and U54-GM094597 to the NeSG (http://www.nesg.org/), a Groupement d′Intérêt Scientifique–CNRS fellowship to S.A., Agence Nationale de la Recherche–Blanc 2010 grant INSERAD and Région Rhône-Alpes grant CIBLE 2008-2011.
PY - 2013/5
Y1 - 2013/5
N2 - How living organisms create carbon-sulfur bonds during the biosynthesis of critical sulfur-containing compounds is still poorly understood. The methylthiotransferases MiaB and RimO catalyze sulfur insertion into tRNAs and ribosomal protein S12, respectively. Both belong to a subgroup of radical-S-adenosylmethionine (radical-SAM) enzymes that bear two [4Fe-4S] clusters. One cluster binds S-adenosylmethionine and generates an Ado• radical via a well-established mechanism. However, the precise role of the second cluster is unclear. For some sulfur-inserting radical-SAM enzymes, this cluster has been proposed to act as a sacrificial source of sulfur for the reaction. In this paper, we report parallel enzymological, spectroscopic and crystallographic investigations of RimO and MiaB, which provide what is to our knowledge the first evidence that these enzymes are true catalysts and support a new sulfation mechanism involving activation of an exogenous sulfur cosubstrate at an exchangeable coordination site on the second cluster, which remains intact during the reaction.
AB - How living organisms create carbon-sulfur bonds during the biosynthesis of critical sulfur-containing compounds is still poorly understood. The methylthiotransferases MiaB and RimO catalyze sulfur insertion into tRNAs and ribosomal protein S12, respectively. Both belong to a subgroup of radical-S-adenosylmethionine (radical-SAM) enzymes that bear two [4Fe-4S] clusters. One cluster binds S-adenosylmethionine and generates an Ado• radical via a well-established mechanism. However, the precise role of the second cluster is unclear. For some sulfur-inserting radical-SAM enzymes, this cluster has been proposed to act as a sacrificial source of sulfur for the reaction. In this paper, we report parallel enzymological, spectroscopic and crystallographic investigations of RimO and MiaB, which provide what is to our knowledge the first evidence that these enzymes are true catalysts and support a new sulfation mechanism involving activation of an exogenous sulfur cosubstrate at an exchangeable coordination site on the second cluster, which remains intact during the reaction.
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U2 - 10.1038/nchembio.1229
DO - 10.1038/nchembio.1229
M3 - Article
C2 - 23542644
AN - SCOPUS:84879097741
SN - 1552-4450
VL - 9
SP - 333
EP - 338
JO - Nature Chemical Biology
JF - Nature Chemical Biology
IS - 5
ER -