Uncovering the molecular strategies that allow human gut symbionts to degrade insoluble dietary and host glycans

Lead Research Organisation: Institute of Food Research
Department Name: Gut Microbes and Health


The human intestine harbours enormous numbers (100 trillion) of resident gut micro-organisms that have important consequences for many aspects of health. The energy sources that support the growth of this complex community derive largely from carbohydrates (glycans) that are not degraded by host enzymes, in particular from dietary plant polysaccharides and host-secreted mucin. Much of the non-digestible carbohydrate that enters the large intestine is in the form of insoluble material such as starch particles, plant cell wall fragments and secreted mucus. Rather few bacterial species have the ability to degrade these insoluble substrates. Those that do must be considered 'keystone species', responsible for releasing energy to the rest of the microbial community, and also, via the uptake of microbially-produced short fatty acid products across the gut wall, providing around 10% of the host's energy supply from the diet. Understanding of microbial glycan utilization is therefore fundamental to understanding the impact of diet upon health, and for developing approaches to manipulate the gut microbiota for health benefit. Almost all of the detailed work so far on glycan metabolism by the human gut microbiota has focussed on gram-negative Bacteroides and there is very little information on the equally numerous gram-positive bacteria belonging to the Firmicutes phylum. Recent evidence indicates however that it is certain Firmicutes, especially Ruminococcus spp., that play 'keystone' roles in initiating the degradation of insoluble substrates, whereas human colonic Bacteroides spp. tend to favour soluble carbohydrates. This proposal will therefore investigate for the first time the molecular mechanisms that enable human colonic species of Ruminococcus to degrade particulate resistant starch (R. bromii), cereal bran rich in plant cell wall polysaccharides (R. champanellensis) and insoluble mucin (R. gnavus). The work will exploit the available genome sequences to enable functional studies on extracellular enzymes, enzyme complexes and substrate attachment mechanisms. Preliminary work already shows that organization at the level of the genome and enzyme systems (including likely extracellular enzyme complexes) is completely different from that in Bacteroides spp. A second main element of the proposal will examine interactions of these primary degraders with other species that are likely to compete for solubilized products of insoluble substrates (including Bacteroides spp.), or to modify metabolism by utilizing fermentation products (acetogenic bacteria). These interactions will be explored in vitro and also in vivo by using gnotobiotic animal models (colonised by single, or combinations of, Ruminococcus strains). The project will substantially advance our understanding of the interdependency of different groups within the human gut microbiota and the impact of variations in gut microbiota composition, and will help to test and predict the fermentability of different types of plant material in the gut. Results from this work will help us understand how to keep a beneficial relationship with our gut bacteria and should lead to the development of novel strategies to maintain a 'healthy' gut microbiota and to re-adjust the microbial community following disturbance ('dysbiosis') eg. caused by antibiotics or disease states.

Technical Summary

The human colonic microbiota gains most its energy by degrading insoluble substrates such as non-digestible plant fibre, starch particles and mucin. Fermentation of these substrates has important consequences for gut metabolism and human health, but is initiated by a few specialised 'keystone' species. This project will investigate for the first time the enzyme systems and attachment mechanisms that enable keystone species of human intestinal ruminococci to degrade insoluble resistant starches (R. bromii), cereal bran (R. champanellensis) and mucin (R. gnavus). This will require functional characterisation of carbohydrate-active systems identified by bioinformatic analysis of draft genomes as playing a key role in the degradation pathways of these insoluble substrates. These include catalytic domains and modules that may be involved in binding to carbohydrate substrates, in protein:protein interactions and in attachment to the bacterial cell surface. In particular the project will investigate the putative roles of dockerin and cohesin modules in assembling enzyme complexes, which are likely to include the first case of an 'amylosome' in any microorganism, and the first case of a 'cellulosome' in a human colonic bacterium; and determine for the first time, the complement of enzymes required for mucin degradation in Firmicutes. The project will also use anaerobic co-culture studies to investigate interactions with other dominant members of the human intestinal microbiota, specifically with hydrogen-utilizing organisms, and with 'secondary' carbohydrate-utilizing species such as Bacteroides thetaiotaomicron. The impact of co-cultures and cross-feeding will be assessed upon bacterial growth, metabolite production and gene expression. Finally, selected co-culture experiments will be translated into gnotobiotic mice in order to relate bacterial utilization of insoluble substrates under in vivo conditions to changes in enzymes and metabolites relevant to health.

Planned Impact

BBSRC Strategy
The research fits with the BBSRC's Bioscience Underpinning Health priority and its Grand Challenge 3 - Fundamental bioscience enhancing lives and improving wellbeing (BBSRC Delivery Plan 2011-15). The work also has potential to benefit the food production and processing sector, and is therefore also relevant to the Food Security Priority.

National Health Service & Consumers
The research has the potential to impact on the nation's health and welfare through reducing the onset and progression of gut-associated diseases. Variation in microbiota composition has been suggested to underlie intolerance of high-fibre diets in certain groups (eg. IBS sufferers) that is associated with excessive fermentation. Irritable bowel syndrome (IBS) is one of the commonest long-term gastrointestinal conditions. It is estimated that 10-20% of the UK's population is affected by IBS at any one time, although this figure may be higher because many people with the condition do not report their symptoms to their GP.
IBS is twice as common in women as it is in men. The condition normally develops in people who are between 20 and 30 years of age, but it can affect people of any age. It is estimated that three out of four people with IBS will have at least one bout of depression, and just over half will develop generalised anxiety disorder (a condition that can cause overwhelming feelings of anxiety, fear and dread). These have a major adverse impact on the potential economic contribution of this demographic.
Prebiotics are selectively fermented, dietary ingredients that result in specific changes in the composition and/or activity of the gastrointestinal microbiota, thus conferring benefit(s) upon host health. Unlike probiotics, a prebiotic targets the microbiota already present within the ecosystem, acting as a 'food' for the target microbes seen as beneficial. To date, the majority of prebiotic strategies have been based on an empirical approach to manipulate levels of bifidobacteria, lactic acid bacteria that make up a small proportion of the adult microbiota. Thus, the potential to specifically manipulate other beneficial members of the gut microbial community remains largely unexplored. The data generated in the project will provide us with the unparalleled opportunity of developing prebiotic strategies, specifically targeted to distinct members of the microbiota, with the aim of modulating levels of metabolites and host-microbe interactions that provide significant health benefits. Such preventive treatments will significantly reduce NHS costs, and improve the health of the nation.

Policy makers & Government
Prebiotics are currently being discussed by working parties of international scientific organizations such as the Food and Agriculture Organization of the WHO and the International Life Sciences Institute and changes to the definition and concept may follow in time. For now, their use as food ingredients or supplements is currently popular and gaining momentum. The research is likely to provide evidence for use of experts who sit on advisory panels that contribute to policy or dietary advice.

Food & Biotech industry,
The outcomes of this project will thus be of direct interest to enzyme companies involved in the development of commercial and non-commercial (e.g. academic enzymes) enzymes. CAZymes are used as biocatalysts in a wide range of industrial biotechnology sectors based on processing of plant and cell wall polysaccharides, encompassing business segments such as Fabric and Household Care (enzymes for laundry and dishwashing detergents), Technical Enzymes (enzymes for carbohydrate processing as well as textile treatment), Pulp and Paper, and Food and Animal Nutrition (enzymes for bread, feed and brewing applications).
Description We have obtained novel mechanistic insights into the ability of some gut bacteria to utilise mucin glycans. We suggested a novel strategy by which gut bacteria adapt to the mucosal environment. Our new data support the hypothesis that mucin-degraders such as Ruminococcus gnavus play a key role in establishing cross-feeding activities to sustain a mucosa-associated microbiota (Crost et al. 2018 Front Microbiol.). In collaboration with Aberdeen we have contributed to unravelling the molecular pathways resulting in resistant starch utilisation in gut bacteria (Laverde Gomez Environ Microbiol. 2019; Mukhopadhya et al., Environ Microbiol. 2018).
Exploitation Route The mechanisms identified during this study provide molecular leads that can be used to establish new strategies to modulate the gut microbiota.
The glycoenzymes characterised as part of this study may have industrial applications in the biotech and biopharmaceutical sectors.
Sectors Agriculture, Food and Drink,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

URL https://quadram.ac.uk/juge-flint-grant/
Description Training interdisciplinary glycoscientists to get a molecular- level grip on glycocodes at the human mucosa-microbiota
Amount £466,300 (GBP)
Funding ID 814102 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 01/2019 
End 12/2022
Description Carbohydrate analysis 
Organisation John Innes Centre
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Supervised and coordinated research project
Collaborator Contribution Provided expertise and access to NMR facility
Impact WO2017134466A1. Patent Application on intramolecular trans-sialidase; Carbohydr Res. 2017 Nov 8;451:110-117. doi: 10.1016/j.carres.2017.08.008.
Start Year 2007
Description Carbohydrate synthesis 
Organisation University of California
Country United States 
Sector Academic/University 
PI Contribution Utilised carbohydrates provided by collaborator in scientific research projects
Collaborator Contribution Provided bespoke carbohydrates upon request
Impact Carbohydr Res. 2017 Nov 8;451:110-117. doi: 10.1016/j.carres.2017.08.008; Nat Commun. 2017 Dec 19;8(1):2196. doi: 10.1038/s41467-017-02109-8.
Start Year 2015
Description Rowett 
Organisation University of Aberdeen
Department Rowett Institute of Nutrition and Health Aberdeen
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided expertise in transcriptomics, metabolomics and access to germ-free mouse facility
Collaborator Contribution Provided resistant starch degrading bacteria
Impact Environ Microbiol. 2018 Jan;20(1):324-336. doi: 10.1111/1462-2920.14000; MBio. 2015 Sep 29;6(5):e01058-15. doi: 10.1128/mBio.01058-15. Food Science & Technology , pp. 24-26
Start Year 2014
Description STD-NMR 
Organisation University of East Anglia
Department Schools of Pharmacy and Medicine UEA
Country United Kingdom 
Sector Academic/University 
PI Contribution We provided proteins and ligands for analysis by STD NMR to complement some of our data on mechanisms of host-bacteria interactions. This collaboration resulted in high impact joint publications led by QIB (Proc Natl Acad Sci U S A. 2018 pii: 201715016; Nat Commun. 2017 ;8(1):2196)
Collaborator Contribution STD- NMR analysis of proteins and ligands we provided. Some of this work led to the development of DEEP STD NMR, leading to a joint publication led by UEA (Angew Chem Int Ed Engl. 2017 56:15289-15293).
Impact Proc Natl Acad Sci U S A. 2018 pii: 201715016; Nat Commun. 2017 ;8(1):2196; Angew Chem Int Ed Engl. 2017 56:15289-15293
Start Year 2014
Description Synchrotron Oxford 
Organisation Diamond Light Source
Country United Kingdom 
Sector Academic/University 
PI Contribution Provided collaborator with proteins and ligands for X-ray crystallography and cryo-EM
Collaborator Contribution X-ray crystallography of proteins free and in complex
Impact Nat Commun. 2017 Dec 19;8(1):2196. doi: 10.1038/s41467-017-02109-8.
Start Year 2016
Description Science Festival in Norwich 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Presentation of an interactive stand and posters at the Science Festival at the Norwich Cathedral on 'Mucus and gut microbes: a sweet relationship to keep us healthy'
Year(s) Of Engagement Activity 2016
Description 4th Annual European Microbiome Congress 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Industry/Business
Results and Impact Attendance to 4th Annual European Microbiome Congress. This Conference has a high representation from the Industry: Pharmaceutical companies and Biotechnology companies and provided an excellent opportunity to make contact and discuss work relevant to the Institute Strategic Programmes. The PROMOTING WOMEN IN SCIENCE lunch session was inspiring.
Year(s) Of Engagement Activity 2018
Description Academia-Industry Networking CarboMet event 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact I co-organised and chaired a two day workshop on 'The Role of Carbohydrates in the Gut Microbiome' June 2018 in Brussels attended by 40 delegates from academia and industry as apart of CarboMet. CarboMet (Metrology of Carbohydrates for European Bioindustries) is a four-year Coordination and Support Action (CSA) funded by Horizon 2020 FET-OPEN. It is an European Network that facilitates engagement between key players and stakeholders of the glycoscience community across Europe to identify the current state of the art and in particular future innovation and technological challenges in carbohydrate metrology. I co-wrote and edited the CarboMet positioning paper on 'The Essential Roles of Carbohydrates in Promoting Gut Microbiota Function Through All Stages of Life' which summarises discussions that took place during the workshop.
Year(s) Of Engagement Activity 2018
URL https://carbomet.eu/news/positioning-paper-on-role-of-carbohydrates-in-gut-microbiota-function/
Description IBCarb 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact Contributed to the final IBCarb workshop Glycobiotechnology 2018. IBCarb (Glycoscience Tools for Biotechnology and Bioenergy) is a growing network of glycoscientists from academia and industry. IBCarb was one of the Networks in Industrial Biotechnology and Bioenergy funded by the BBSRC. The workshop was very successful and attracted around 150 attendees (international audience from academia and industry) . This was an opportunity to showcase the Glycosciences carried out in the UK and in partnership with industrial/business partners.
Year(s) Of Engagement Activity 2018
URL https://ibcarb.com/events/