Revealing glycosylation patterns of mucins.

Project: PhD

Project Details


The condition of the intestine mucus is of essence to maintain a healthy gut and a protective barrier function. Due to the complexity of the mucus structure and the quantitative and qualitative analysis of glycans in mucins, there is a lack of understanding regarding the influence of mucin glycosylation on gut health and vice versa. Specifically, which mechanisms are utilized by certain gut microbes in relation to mucin glycosylation and how mucin glycosylation is shaped as a result. It is suggested that specific gut microbes have a positive effect on, for example, mucin gene expression and mucus growth and that dietary fibre has a positive effect on mucus health. Part of this research will therefore include complete characterization and (semi) quantification of O-linked and N-linked glycans, including sialylated glycans, present in mucins extracted from in vitro mucus expressed by HT29-MTX-E12 cells. To understand the interaction between gut conditions and mucus, these cells will be exposed to pasteurized bacteria such as A. muciniphila, and pre-biotics such as dietary fibres[1] [28]. This will give significant insight in the changes in mucin glycosylation in relation to the condition and variations of the gut microbiome. We expect to characterise a wide variety of glycans in the in-vitro mucus samples (especially O-glycans). Furthermore, we expect that a divers gut microbial community as well as a fibre-rich diet stimulate the mucus layer to grow thick and feature a larger variety of glycans. Certain ‘’types’’ of glycan structures might be more abundant than others. As microbes and glycans interact with each other, we expect the variety of glycans to stimulate the diversity of the gut microbiome. We also think that certain glycans or types of glycan structures are indicative for gut health (either positive or negative) and that the abundance of glycans can support the understanding of this relationship. The key objectives are: 1. Setting up a mass spectrometric analysis method to fully characterise and (semi) quantify released O-linked and N-linked glycans (including sialic acids) using commercial standards and in vitro produced mucus material (including exposure to B. fragilis, R. gnavus, and A. muciniphila) 2. Investigating how specific gut microbes influence the variety and abundance of O-linked and N-linked glycans using in vitro produced mucus material (resembling intestine mucus). In addition, establishing which enzymes are involved in the process and which sugars and glycosyl linkages are addressed during enzymatic modifications. Furthermore, mapping enzyme specificity for specific sugar moieties or linkages. 3. Investigating in what way pre-biotics and dietary fibres can influence the intestine mucins/mucus by analysing the changes in amount and position of specific sugar building blocks within mucus glycans and the position of specific glycans within the protein backbone using in vitro produced mucus material. Lastly, exploring the impact on the glycans when part of the building blocks are removed by microbes, for example. 4. Evaluating the previous findings using in vivo gut mucus (obtained via Wilma Steegenga or Clara Belzer or others)
Effective start/end date1/09/21 → …


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