Part I: Crystallization of A Type IV Pilin from Pseudomonas Aeruginosa. Part II: Characterization of a Peptidyl-Prolyl-Cis,Trans-Isomerase Through X-Ray Crystallography

Within a host, pathogenic bacteria employ several mechanisms that enhance their survival and motility. These mechanisms include their ability to adhere, replicate (in order to avoid eradication), and secretion of virulent proteins. The study of proteins involved in bacterial pathogenesis provides us with a more thorough understanding of their mechanism and function, which can lead to the development of more effective therapeutics. In this study proteins secreted by gram-negative bacteria are explored, specifically those from Pseudomonas aeruginosa and Helicobacter pylori. P. aeruginosa is a common opportunistic pathogen associated with 10% of hospital infections, mainly owed to their ability to bind to biotic and abiotic surfaces. Type IV pili secreted by P. aeruginosa are associated with adhesion, motility and DNA transfer. Part I details the cloning to crystallization of KB7, a pilin secreted by P. aeruginosa. The second bacteria studied in Part II is H. pylori, associated with gastric ulcers and gastric inflammation compromising 50% of the global population, where severity of infection is highly dependent on the strain of H. pylori, and the individual infected. The secreted protein HP0175 from H. pylori binds to Toll-Like Receptor 4 and activates a cascade of mechanisms leading to apoptosis, as well as triggering the innate immune response. HP0175 is also classified as a peptidyl-prolyl cis,trans-isomerase involved in the isomerization of proline peptide bonds preceding the N-terminal. Here, apo-HP0175 was crystallized to 2.09 in space group P3221 with one monomer in the asymmetric unit; the dimer is generated through symmetry mates. A comparison to indole-2-carboxylic acid bound HP0175 shows N- and C- terminal helix extensions upon interaction of the catalytic residues in the binding pocket. Helix extension supports other parvulin findings that N- and C- terminal helices stabilize proteins undergoing catalysis by protein-protein interaction