Biomol NMR Assign (2011) 5:35–38 DOI 10.1007/s12104-010-9261-6 A R T I C L E NMR resonance assignments of NarE, a putative ADP-ribosylating toxin from Neisseria meningitidis Ludovic Carlier • Christian Koehler • Daniele Veggi • Mariagrazia Pizza • Marco Soriani • Rolf Boelens • Alexandre M. J. J. Bonvin Received: 24 June 2010 / Accepted: 13 August 2010 / Published online: 25 August 2010 The Author(s) 2010. This article is published with open access at Springerlink.com Abstract NarE is a 16 kDa protein identiﬁed from Neisseria meningitidis, one of the bacterial patho- gens responsible for meningitis. NarE belongs to the ADP-ribosyltransferase family and catalyses the transfer of ADP-ribose moieties to arginine residues in target protein acceptors. Many pathogenic bacteria utilize ADP-ribosy- lating toxins to modify and alter essential functions of eukaryotic cells. NarE was proposed to bind iron through a Fe–S center which is supposed to be implied in catalysis. We have produced and puriﬁed uniformly labeled 15N- and 15N/13C-NarE and assigned backbone and side-chain res- onances using multidimensional heteronuclear NMR spectroscopy. These assignments provide the starting point for the three-dimensional structure determination of NarE and the characterization of the role of the Fe–S center in the catalytic mechanism. Keywords ribosylation NarE Pathogenic bacteria Meningitis NMR ADP L. Carlier C. Koehler R. Boelens A. M. J. J. Bonvin (&) Bijvoet Center for Biomolecular Research, Science Faculty, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands e-mail: a.m.j.j.bonvin@uu.nl D. Veggi M. Pizza M. Soriani Novartis Vaccines and Diagnostics, 53100 Siena, Italy Present Address: L. Carlier Laboratoire des Biomole´cules, CNRS UMR7203, UPMC Universite´ Paris 06, 75005 Paris, France Biological context Neisseria meningitidis is a gram-negative bacterium best known for its role in meningitis and septicemia. This human-speciﬁc pathogen is considered as a major cause of morbidity and mortality during childhood in industrialized countries and is responsible for epidemics in Africa and in Asia (Genco and Wetzler 2010). Like most bacterial pathogens, N. meningitidis synthesizes a number of toxic products believed to target and kill their host cells. Among these toxins, proteins that exert ADP-ribosylation activity represent a large family of potentially lethal enzymes able to modify or disrupt essential functions of eukaryotic cells (Moss and Vaughan 1988). ADP-ribosyltransferases cata- lyze the transfer of a single ADP-ribosyl group from b-nicotinamide adenine dinucleotide (NAD?) onto speciﬁc amino acid residues of host cell proteins with the simul- taneous release of nicotinamide (Althaus and Richter 1987). Many bacterial pathogens use ADP-ribosylating enzymes to block protein synthesis or to alter signal transduction by inactivating key target proteins such as GTP-binding proteins (Locht and Keith 1986). The putative ADP-ribosylating NarE protein from N. meningitidis was recently identiﬁed on the basis of a proﬁle-based computational approach (Masignani et al. 2003). NarE shares structural features with toxins from V. cholerae and E. coli, and, like cholera toxin and heat-labile enterotoxins, retains the capacity to ADP-ribosylate argi- nine and to hydrolyze NAD in ADP-ribose and nicotin- amide. Another feature of NarE is its ability to bind iron through an iron–sulfur center (Fe–S) (Del Vecchio et al. 2009). Interestingly, site-directed mutagenesis and enzy- matic assays showed that correct assembly of the iron- binding site is essential for transferase but not for hydrolase activity (Del Vecchio et al. 2009), suggesting for the ﬁrst 123