RESEARCH ARTICLE The ROP2 family of Toxoplasma gondii rhoptry proteins: Proteomic and genomic characterization and molecular modeling Hiba El Hajj 1 , Emmanuelle Demey 2 , Joël Poncet 2 , Maryse Lebrun 1 , Bo Wu 3 , Nathalie Galéotti 2 , Marie Noëlle Fourmaux 4 , Odile Mercereau-Puijalon 5 , Henri Vial 1 , Gilles Labesse 6, 7 and Jean François Dubremetz 1 1 UMR 5539 CNRS, Université de Montpellier 2, Place E. Bataillon, Montpellier, France 2 Institut de Génomique Fonctionnelle, UMR 5203 CNRS, U661 INSERM, Université de Montpellier 1, Université de Montpellier 2, Montpellier, France 3 Department of Biological Sciences, University of Pennsylvania, Goddard Laboratories, Philadelphia, PA, USA 4 AD INSERM 7, Place de Verdun, Lille, France 5 Unité d’Immunologie Moléculaire des Parasites, Institut Pasteur, Paris, France 6 CNRS, UMR 5048/Centre de Biochimie Structurale/Université de Montpellier 1, Montpellier, Cedex, France 7 INSERM, U554/Centre de Biochimie Structurale, Montpellier Cedex, France Four rhoptry proteins (ROP) of Toxoplasma gondii previously identified with mAb have been af- finity purified and analyzed by MS; the data obtained allowed the genomic sequences to be assigned to these proteins. As previously suggested for some of them by antibody crossreactivity, these proteins were shown to belong to a family, the prototype of which being ROP2. We describe here the proteins ROP2, 4, 5, and 7. These four proteins correspond to the most abundant prod- ucts of a gene family that comprises several members which we have identified in genomic and EST libraries. Eight additional sequences were found and we have cloned four of them. All members of the ROP2 family contain a protein-kinase-like domain, but only some of them pos- sess a bona fide kinase catalytic site. Molecular modeling of the kinase domain demonstrates the conservation of residues critical for the stabilization of the protein-kinase fold, especially within a hydrophobic segment described so far as transmembrane and which appears as an helix buried inside the protein. The concomitant synthesis of these ROPs by T. gondii tachyzoites suggests a specific role for each of these proteins, especially in the early interaction with the host cell upon invasion. Received: March 16, 2006 Revised: June 20, 2006 Accepted: June 20, 2006 Keywords: Protein kinase / Protein structure prediction / Rhoptry / Toxoplasma Proteomics 2006, 6, 5773–5784 5773 1 Introduction The protozoan phylum Apicomplexa includes a large num- ber of human and animal parasites responsible for a wide variety of diseases. Plasmodium sp. are the causative agents of malaria, Toxoplasma gondii is the agent of toxoplasmosis and the other apicomplexan parasites such as Cryptosporidium Correspondence: Dr. Jean François Dubremetz, UMR 5539 CNRS, CC 107, Université de Montpellier 2, Place E. Bataillon, 34095 Montpellier, France E-mail: dubremet@univ-montp2.fr Fax: 133-467144-286 Abbreviations: PV, parasitophorous vacuole; PVM, PV mem- brane; ROP , rhoptry protein DOI 10.1002/pmic.200600187  2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.proteomics-journal.com