Structure and function of the oligopeptide permease transport system in the cell wall of Moraxella catarrhalis, a human respiratory pathogen
Jones, Megan Makenzie
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Moraxella catarrhalis is a strict human pathogen that contributes to worldwide morbidity and mortality by causing otitis media in children and exacerbations of chronic obstructive pulmonary disease in adults, the third leading cause of death in the United States. Regarded as a commensal organism for many years, M. catarrhalis is an understudied pathogen with little understood about its metabolic pathways or mechanisms of virulence. Vaccine development is a priority for M. catarrhalis and the OppA protein is a potential antigen candidate. The oppA gene is the predicted substrate binding protein of the putative oligopeptide permease ABC transport system ( opp ) consisting of 5 genes ( oppB, C, D, F , and A ), encoding 2 permeases, 2 ATPases, and a substrate binding protein. These five opp genes are in the same open reading frame that contains a second promoter within the intergenic region of sequence between oppF and oppA. In gram-negative bacteria, substrate binding proteins are usually found in the periplasm, with permease and ATPase proteins forming a pore complex in the cytoplasmic membrane. M. catarrhalis has a growth requirement for arginine, thus acquiring arginine is important for fitness and survival. We hypothesized that the putative Opp system plays an important role in fitness through peptide uptake for arginine acquisition and may be regulated by the temperature and/or nutrient availability. We showed that the gene cluster was transcribed as an operon with two functional promoters. OppA was found to bind peptide substrates ranging from 3 amino acid residues to at least 16 amino acid residues in length, while the size of peptide imported by the entire Opp system was limited to 5 to 10 amino acid residues in length. We also showed M. catarrhalis requires L-arginine and cannot utilize methylated arginine, which is abundant in the respiratory tract. Cold shock at 26 °C for ≤ 0.5 hours and the presence of a peptide substrate increases all opp gene transcript levels. Furthermore, an oppA mutant showed marked impairment in its capacity to persist in the murine respiratory tract compared to wild type. We show that OppA is located in multiple locations in the bacterial cell wall and also binds to vitronectin, a ubiquitous human matrix protein. This indicates a possible role of OppA in adhesion and colonization of the human respiratory tract. We conclude that the Opp system is an important nutritional virulence factor regulated by temperature and nutrient availability in M. catarrhalis and that OppA shows potential as a vaccine antigen. The Opp system represents a possible target for new antimicrobial agents and the development of OppA as a vaccine antigen may not only yield protective antibodies, but also antibodies that could interfere with virulence. The work presented in this thesis advances the field by elucidating the role of the Opp system in fitness and virulence of M. catarrhalis , contributing to the understanding of how this important human pathogen operates in the harsh environment of the human respiratory tract.