Identification and Characterization of the Moraxella catarrhalis IgD-binding Protein MID

Abstract: Moraxella catarrhalis is now an established human pathogen that frequently colonizes the upper respiratory tract causing most importantly otitis media in children. It has also been associated with sinusitis, laryngitis and lower respiratory tract infections in adults with predisposing conditions such as chronic obstructive pulmonary disease (COPD). Current research mainly focuses on surface antigenic determinants of M. catarrhalis as it progresses towards a future vaccine. MID is an adhesin that binds both soluble and membrane-bound IgD in a non-immune fashion. The membrane protein MID was isolated by solubilization using the zwitterionic detergent Empigen and then purified by ion exchange chromatography and gel filtration. The 2139 amino acids long protein ? as deduced from the cloned mid gene ? behaved as a multimer under native conditions. We investigated the prevalence of MID in 91 clinical isolates and 7 culture collection strains. The conserved mid gene was found in all strains, however the protein expression varied independently of anatomical site of isolation or geographical origin of the strains. We also showed that the mid gene was regulated by phase variation controlled by a poly(G) box within the open reading frame. Mapping the membrane-bound MID with gold-labeled specific antibodies in transmission electron microscopy (TEM) revealed a double-folded 100 nm long fibrillar structure with a globular distal domain responsible for both the IgD-binding and adhesive capacities of MID. In addition, we demonstrated that MID is a multimer and consists of trimers, tetramers and pentamers. Furthermore, sequence analysis also showed that MID is an autotransporter that belongs to the Oligomeric coiled-coil adhesin (Oca) family. Finally, the human erythrocyte membrane protein adducin was identified as a putative receptor for the adhesive domain MID764-913. This is the underlying interaction for the hemaglutinating activity of M. catarrhalis. The isolation and characterization of a major novel outer membrane protein designated MID (Moraxella IgD-binding protein) has been a significant contribution to the field. The papers in this thesis describe the prevalence, gene regulation, domain structural organization and receptor specificity of MID.