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

人体肠道内有大量（100万亿）常驻肠道微生物，对健康的许多方面都有重要影响。支持这种复杂群落生长的能量来源主要来自不被宿主酶降解的碳水化合物（聚糖），特别是来自膳食植物多糖和宿主分泌的粘蛋白。进入大肠的许多不易消化的碳水化合物是以不溶性物质的形式存在的，如淀粉颗粒、植物细胞壁碎片和分泌的粘液。相当少的细菌物种具有降解这些不溶性底物的能力。那些这样做的必须被认为是“关键物种”，负责向微生物群落的其余部分释放能量，并且还通过肠道壁吸收微生物产生的短脂肪酸产物，提供约10%的宿主饮食能量供应。因此，了解微生物聚糖的利用对于了解饮食对健康的影响以及开发操纵肠道微生物群以获得健康益处的方法至关重要。到目前为止，几乎所有关于人类肠道微生物群聚糖代谢的详细工作都集中在革兰氏阴性拟杆菌上，而关于同样数量的属于厚壁菌门的革兰氏阳性细菌的信息很少。然而，最近的证据表明，某些厚壁菌门，特别是瘤胃球菌属，在启动不溶性底物的降解中起“关键”作用，而人结肠拟杆菌属（Bacteroides spp.）倾向于可溶性碳水化合物。因此，这项提议将首次研究使人结肠瘤胃球菌降解颗粒抗性淀粉的分子机制（R。bromii）、富含植物细胞壁多糖的谷物麸皮（R. champanellensis）和不溶性粘蛋白（R. gnavus）。这项工作将利用现有的基因组序列，使胞外酶，酶复合物和底物附着机制的功能研究。初步工作已经表明，在基因组和酶系统（包括可能的胞外酶复合物）水平上的组织与拟杆菌属完全不同。该提案的第二个主要内容将检查这些主要降解物与可能竞争不溶性底物的溶解产物的其他物种（包括拟杆菌属）的相互作用，或通过利用发酵产物（产乙酸细菌）来改变代谢。这些相互作用将在体外和体内通过使用gnotobiotic动物模型（由单一或组合的瘤胃球菌菌株定殖）进行探索。该项目将大大推进我们对人类肠道微生物群中不同群体相互依赖性的理解，以及肠道微生物群组成变化的影响，并将有助于测试和预测肠道中不同类型植物材料的发酵能力。这项工作的结果将帮助我们了解如何与我们的肠道细菌保持有益的关系，并应导致开发新的策略来维持“健康”的肠道微生物群，并在干扰（“生态失调”）后重新调整微生物群落。由抗生素或疾病状态引起。

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.