Molecular Mechanisms of the Type I Secretion System from Bacterial Pathogens

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ABSTRACT Protein secretion is an essential component of the arsenal used by many bacterial pathogens to invade hosts, damage tissues, and suppress immune responses. Gram-negative bacteria have evolved sophisticated machines to secrete proteins across two membranes: the inner membrane and outer membrane. Since discovered in 1979, the type I secretion system (T1SS) has been identified in more than 800 bacterial genomes, and many of its substrates contribute to the virulence of pathogens, examples being hemolysin HlyA in uropathogenic Escherichia coli, adhesin SiiE in Salmonella enterica, and adenylate cyclase CyaA in Bordetella pertussis. The T1SS secretes unfolded substrates from the cytoplasm to the extracellular milieu. It includes three components: an ATP-binding cassette transporter (ABC transporter), associated periplasmic adapter proteins located in the inner membrane, and a porin in the outer membrane. The canonical “alternating access” model cannot explain how the large protein substrates are transported by the T1SS ABC transporter across the inner membrane. Therefore, the mechanisms of substrate secretion at the molecular level is still obscure. The proposed research leverages single particle cryo-electron microscopy (cryoEM), bioinformatics, in vivo and in vitro assays to answer key questions in the field, including the structural basis of substrate recognition, the nature of the translocation pathway, the energetics for substrate translocation, and the regulation of the T1SS ABC transporter's activity. The studies will substantially increase the fundamental scientific knowledge about the mechanisms of the T1SS and provide a new basis for developing future therapeutic interventions against a broad range of bacterial infections.