Structural analysis of the Escherichia coli β-barrel assembly machinery complex

The outer membrane (OM) is a unique structural feature of Gram-negative bacteria. Residing within the outer membrane are β-barrel outer membrane proteins (OMPs) that serve many important cellular functions. As proper folding and assembly of these proteins are crucial for cell viability, Gram-negative bacteria possess a specialized proteinaceous machine, known as the BAM (β-barrel assembly machinery) complex, to catalyze the folding and membrane insertion of OMPs. In Escherichia coli, the BAM complex consists of five proteins: one β-barrel membrane protein – BamA, and four lipoproteins – BamB, BamC, BamD, and BamE. The roles of the individual components and how they are arranged into the BAM complex to function together is not yet clearly understood. During the course of this thesis project, I determined the structures of E. coli BamB, BamC, BamE and the BamCD subcomplex. Analysis of the conserved residues and the molecular surface properties of these solved structures helped to identify potential protein-protein interaction sites on each lipoprotein. For example, BamC has two ‘helix-grip’ domains that are ideally shaped to accommodate α-helices. BamB, on the other hand, has a β-propeller fold that could potentially interact with BamA or substrates via β-augmentation, a mode of interaction in which a pre-existing β-sheet is augmented by an addition of a β-strand of another protein. Comparing the solved structures with their structural homologs with known functions has also provided important clues about the functional roles of each lipoprotein. BamD structure, for example, closely resembles the binding pocket of a peroxisomal targeting signal receptor PEX5, suggesting a similar substrate recognition function for BamD. Interestingly, our BamCD complex structure shows that the putative substrate binding pocket of BamD is bound and blocked by the conserved unstructured N-terminal region of BamC. This suggests a possibility that BamC may have a regulatory function.The structural and interaction data acquired from this thesis project contributes to a better understanding of the BAM complex structure and provides a platform for future research driven by structure-based hypotheses