Claudio Piselli

• During my Ph.D. I focused on the characterization of the biological membrane channels that are involved or interfering with the communications between the cells and the environment, or the rest of the organism. My thesis is divided in 3 parts: outer membrane proteins, cell wall channels and toxins. All the pore forming proteins were studied in electrophysiological experiments via their reconstitution into the artificial lipid bilayer. 1 In the first part we studied the transport of ions and antibiotics through the phosphate specific channels OprO and OprP expressed in the outer membrane of Pseudomonas aeruginosa. Fosmidomycin and fosfomycin are phosphonic acid containing antibiotics that we kept in consideration for our studies; they are drugs approved from the FDA very active against Gram-positive and Gram-negative bacteria and even protozoa. Then we produced and analyzed mutants of OprO affecting the lysine cluster in the periplasmic cavity of each OprO monomer. The molecular details of these mechanisms are elucidated via the computational analysis performed by Dr. Vinaya Kumar Golla from the workgroup of Professor Ulrich Kleinekathöfer. 2 In the second part we studied the cell wall channels expressed in Rhodococcus ruber and corynebacterioides. Like for all the MspA homologs, the PorARr, PorBRr, PorARc are synthetized as monomers and are able to form octamers in their active form as channels in which they can allow the passage of nutrients and secondary metabolites, such as antibiotics, driven by concentration gradient. 3 In the third part we studied 3 pore forming toxins (PFTs): the ε-Toxin and β2-Toxin from Clostridium perfringens and the LktA of Mannheimia haemolytica. Bacteria can finalize their virulence through PFTs that make large water-filled holes into the target membrane dissipating its osmotic gradient.