Structural basis for nutrient acquisition by dominant members of the gut microbiota

The large bowel is colonised by an extremely dense population of bacteria, collectively termed the colonic microbiota. Recent research indicates that the microbiota play an important role in maintaining normal health and nutrition. Imbalances in the microbiota have been linked to a wide range of diseases from autoimmune disease to cancer and obesity. The function and composition of the microbiota is dependent on the ability of individual micro-organisms to acquire nutrients such as starch and other dietary polysaccharides in the highly competitive environment of the human large bowel. This process of nutrient acquisition is carried out by protein machines embedded in the bacterial cell envelope. In many microbiota members, this machine is a two-component complex consisting of a substrate binding protein (termed SusD) and a channel-forming transport protein (termed SusC). We have determined the first three-dimensional atomic structure of a SusCD complex by X-ray crystallography and have obtained good-quality crystals for another SusCD complex. These exciting preliminary data form the basis of this proposal, in which we will elucidate in detail how these SusCD complexes transport their substrates into the bacterial cell. By answering a number of "how does it happen?" questions, our project will provide fundamental insights into the functioning of the microbiota and understanding the human-microbiota symbiosis. Importantly, such results will be a timely and necessary complement to most current microbiota research, which is focused on answering questions such as "who is there and when?". By linking mechanistic and systems biology, our project could also have practical implications by providing insights to manipulate the composition of the microbiota via interference with critical nutrient uptake processes.

The human large intestine is populated by an extremely high density of microorganisms, collectively termed the colonic microbiota. It is becoming increasingly evident that the microbiota has a large influence on human health and nutrition. The composition and therefore the collective function of the colonic microbiota depends on the ability of individual micro-organisms to acquire nutrients in the highly competitive environment of the large intestine. The two dominant phyla are the Gram-positive Firmicutes and the Gram-negative Bacteroidetes, in particular the genus Bacteroides. Contrasting with the small intestine where small-molecule nutrients such as simple sugars and amino acids are abundant, the major source of nutrients for the colonic microbiota are polymeric glycans derived from e.g. dietary fiber. In Bacteroides spp., those glycans are processed by surface enzymes into oligosaccharides and imported by an outer membrane (OM) protein complex consisting of an extracellular SusD-like substrate binding lipoprotein and a SusC-like integral membrane transporter. Due to a lack of structural information, it is not known how this crucial process occurs. In an important step towards this goal we have purified a SusCD complex from Bacteroides thetaiotaomicron and determined its X-ray crystal structure to 2.75 Å resolution. The structure shows a number of novel and unexpected features and forms the basis for this proposal, in which we will elucidate in detail how SusCD complexes generate and select their substrates and how these are subsequently transported across the OM into the cell. Given the abundance and importance of SusCD systems for the functioning of Bacteroidetes, our project will provide a major advance in the molecular-level understanding of the microbiota and provides a timely and necessary complement to a multitude of recent high-profile (meta)genomics studies.

Our gut microbiota play a pivotal role in maintaining health and nutrition and this is increasingly being recognised by the food industry who are very keen to understand mechanisms by which we can modulate the composition of our microbiota to improve human health through dietary-based prebiotic and nutraceutical strategies. Key to the development of these dietary strategies is a mechanistic understanding of nutrient acquisition by the gut microbiota. This proposal seeks to provide this insight and thus the underpinning knowledge that allows us to design specific molecules to modulate microbiota function.

The huge amount of sequence data generated by the recent Human Microbiome and MetaHit projects has provided a significant insight into the identity of the key members of the microbiota and the metabolic traits they encode. However, these large scale sequencing projects have also highlighted how little mechanistic understanding we have of the drivers of microbial survival in the gut. Imbalances in the microbiota have been implicated in a range of serious disease states, from inflammatory bowel disease to the alarming rise in 'Western world' illnesses such as cardiovascular disease and diabetes. Thus, functional studies on the microbiota, proposed here, will provide much needed insight into the mechanisms underpinning a healthy gut microbiota and will be of interest to both health professionals and medical researchers seeking to manipulate imbalanced microbiota to return them to a healthy state.