M. Nálezková1, L. Blanchard2, A. de Groot2, P. Gans2, D. Marion2


1National Centre for Biomolecular Research, PřF MU, Kotlářská 2, 61137 Brno, Czech Republic, 2L’Institut de Biologie Structural, 41, rue Jules Horowitz, 38027 Grenoble, France


            The analysis of the hyperthermophilic archaeon Pyrococcus abyssi genome showed that some proteins are highly conserved between Archaea and Eukaryotes. 32 proteins have been listed and called the PACE proteins (Proteins of Archaea Conserved in Eukaryotes) [1]. One of these proteins, PACE11, we decided to study by NMR. This protein has been biochemically characterized as a monomeric phosphopantetheine adenylyltransferase (PPAT) involved in the Coenzyme A (CoA) biosynthesis pathway [2]. This pathway, consisting of 5 steps, has been described in Bacteria and Eukaryotes but not in Archaea. PPAT is involved in 4th step of the CoA biosynthesis, catalyzing the reversible transfer of an adenylyl group from ATP to 4‘-phosphopantetheine giving dephosphoCoA (dPCoA). So far the 3D structure of only a bacterial PPAT (from E. coli and T. Thermophilus) has been solved [3]. PACE11 shows very low sequence identity with these bacterial proteins.

            PACE11 project is to get structural and dynamical information of this protein. PACE11 is PPAT of Pyrococcus abyssi, which is composed of 157 residues. In order to get enough protein for the NMR study a synthetic gene of PACE11 (having no E. coli rare codons) was cloned into the pET28a plasmid and the expression of the gene was tested showing a very good production of PACE11. A 15N sample was then prepared and the 1H-15N HSQC recorded on this sample showed that PACE11 is well folded.


[1]  Matte-Tailliez O, Zivanovic Y, Forterre P. (2000), Trends Genet 12, 533-536

[2]  Armengaud J, Fernandez B, Chaumont V, Rollin-Genetet F, Finet S, Marchetti C, Myllykallio H, Vidaud C,          Pellequer JL, Gribaldo S, Forterre P, Gans P. (2003), J Biol Chem 278, 31078-31087

[3]  Izard T. (2002), J Mol biol 315, 487-495