Novartis Vaccines Research Centre, Italy
Ilaria Ferlenghi has received her Ph.D in Biophysics at the University of Heidelberg, Germany, working on viral structures with the pioneer of the cryo-electron microscopy, Dr. Stephen Fuller. She performed several post-doctoral studies at different institutions: at the EMBL, Germany, with Dr S. Fuller and Dr. S.J. Butcher, at the CNRS (France) with Dr. Felix A. Ray applying x-ray crystallography on viral protein structures and later on with Dr. M. Knossow on bacterial helicases structures. During the post-doctoral period she was visiting the laboratory of Dr. Stephen Harrison at the Harvard Medical School, Cambridge, US. She became head of the Structural Microscopy Group at the Novartis Vaccines in the 2006. She has been appointed PhD Director of the Novartis Academy in 2012. She is member of the WILS (Women in Life Science). She has published several book chapters and more than 20 peer-reviewed research papers in reputed international journals. Her research interests focus on the use of structural biology applied to vaccine discovery.
Alpha-hemolysin (Hla), is a major cytotoxic agent released by Staphylococcus aureus and the first identified member of the pore forming beta-barrel toxin family. The S.aureusHla protein monomer assembles into heptameric pores on eukaryotic cellular membrane during bacterial infection causing lysis, apoptosis and junction disruption. The proposed pore forming mechanism consists in a three steps model according to which the Hlaprotomer binds the membrane, forms a heptamericprepore structure and finally rearranges itself into the complete pore structure. Herein we present the design and the three dimensional structure obtained by single-particle reconstruction of a newly engineered S. aureus alpha-toxin HlaPSGS, a recombinant protein devoid of the portion responsible for toxic activity. The structure, determined to medium resolution, maintains the homo-oligomericheptameric assembly while missing the solvent-filled channel present in the Hlawt. This protein demonstrates conclusively the three step mechanism as it retains the wild type overall conformation and polymerization capabilities, forming complete heptameric pre-pore structures but lacks the hemolytic competency of the complete pore structure. Furthermore we demonstrate that the portion responsible for toxic activity is not essential for membrane binding or for the formation of an oligomericprepore intermediate. The HlaPSGSheptameric structure provides insight into the principles of membrane interaction and transport activity of beta-barrel pore-forming toxins. Finally, the absence of cytotoxicity makes this molecule a safe vaccine candidate.