Structural Studies into E. coli SslE and its role in Diarrhoeal Diseases

Bacteria are single cell organisms that inhabit a wide range of environments and can have both a positive and negative impact on human health. The gastrointestinal tract in humans is covered by a layer of mucus and this acts as a barrier to separate host cells from "good bacteria" but also from invading pathogens. However, dangerous bacteria have developed ways to bind and break down host mucus, which releases nutrients to help the bacteria grow but also allows them to access host tissues so they can bind and initiate infection. In this application, we propose a number of experiments that will help us to understand how some dangerous bacteria are able to cause disease in humans through interactions with mucus. We will specifically be studying Escherichia coli, which is commonly found in the human intestines and is often beneficial to the host. However, infectious strains cause severe diarrhoeal diseases, urinary tract infections, blood infections (sepsis) and meningitis. For example, enterotoxigenic E. coli (ETEC) is a strain which is responsible for up to 400 million cases of diarrhoea each year in the developing world, results in the death of at least 300,000 children and is the most frequent cause of traveller's diarrhoea.

E. coli use a 'type II secretion system' (T2SS), a syringe-like mechanism to transport proteins into their surroundings and many of these secreted proteins help the bacteria to cause disease. One such protein, SslE, is secreted by many infectious and some beneficial E. coli strains. Once exported it can break down mucus in the intestines but also allows E. coli to form biofilms, where bacteria clump together and are protected from the local environment. In pathogenic strains, SslE has a major role in promoting bacterial disease. We have been studying SslE from both a harmless and an enterotoxigenic strain and we have begun to unravel how SslE is able to perform these two different functions. Understanding the details of how this protein functions and how it interacts with host mucus will be crucial to further our knowledge of E. coli infection in diarrhoeal and other diseases. Furthermore, this secreted protein has shown great promise as a target to develop new vaccines for a range of different types of disease-causing E. coli strains, so providing a better understanding here will also aid future vaccine development. These studies may also reveal common pathways for infectious disease used by other bacteria, which may in turn help us design compounds which disarm E. coli and other dangerous pathogens.

The gastrointestinal tract is populated by a complex community of microorganisms, the gut microbiota, which have a major role in maintaining a healthy immune system. E. coli is a gram-negative bacterium that colonizes the human intestine and is generally beneficial; however, virulent strains are the cause of enteric diseases, urinary tract infections, sepsis and meningitis. The epithelial cells that line the gastrointestinal tract are covered by a layer of secreted mucus which acts as a barrier to separate epithelial cells and adjacent host tissue from the residing commensal microbiota but also invading pathogens. Mucins are a family of large glycosylated proteins and the primary constituents of mucus. As such, a major pathogenic mechanism of successful enteric pathogens is the targeting and degradation of these mucin structures.
A broad range of E. coli pathotypes export the protein SslE, which is essential for intestinal colonization and has a dual role in promoting mucosal clearance in the host intestine and promoting biofilm maturation. However, we lack a full structural and mechanistic understanding of how SslE interacts with mucins. We have identified a unique feature of SslE where it can form polymers that stabilize biofilm structures and our preliminary studies have provided a partial Cryo-EM structure of the monomeric SslE core region which hints at a novel mode of interaction with mucins. Using the commensal E. coli W and enterotoxigenic H10407 strains as model systems, we now plan to (i) elucidate the atomic structure of intact SslE, (ii) determine how SslE recognizes and processes mucins and (iii) establish the role of SslE/mucin interactions during colonization. These studies will provide insight into new mechanisms related to E. coli pathogenesis and may have implications for the development of new vaccines and strategies to reduce bacterial virulence.