Molecular Characterization and Functional Analysis of the Pilq_380-706: A Novel Secretin Domain in Pseudomonas Aeruginosa

Background: type 4 pili (t4p) is an important virulence factor of pseudomonas aeruginosa (p. aeruginosa). t4p pass the outer membrane through a large oligomeric channel made of a single pilq protein that is most highly conserved at their c-termini. to develop a functional vaccine that can be used in clinical application, the secretin domain of the pilq (pilq380-706) was produced as a recombinant protein. methods: a 981 bp fragment of c-terminal of the pilq secretin (pilq1138-2118) from was designed into the prokaryotic expression vector pet28a. the presence of the pilq1138-2118 gene in the recombinant construct (pet28a/pilq) was assessed by double digestion and pcr. after transformation, expression of the recombinant pilq was induced by addition of iptg. the expressed recombinant protein was purified by a modified method using a histrap affinity column and finally confirmed by sds-page. the functional activities of the produced pilq380-706 confirmed by western blot analysis and twitching inhibition assay. results: the pcr and enzymatic digestion results showed the presence of the pilq1138- 2118 gene in the construct. the protein electrophoresis showed that the molecular weight of the recombinant pilq380-706 is approximately 37 kda. the western blot analysis confirmed the specificity of specific igg against the pilq380-706 protein. the pilq380-706 protein showed high biological activity in all of these standard assays. conclusion: since, the pilq380-706 protein plays an important role in the biogenesis of pili; and thus, the primary establishment of p. aeruginosa; it seems that it can be used as a candidate vaccine or an adjuvant in the future studies.