How sweet are our gut beneficial microbes: Protein glycosylation in Lactobacillus reuteri

Latousakis, Dimitrios (2017) How sweet are our gut beneficial microbes: Protein glycosylation in Lactobacillus reuteri. Doctoral thesis, University of East Anglia.

[thumbnail of Dimitrios_Latousakis_PhD_Thesis_2017.pdf]
Download (7MB) | Preview


Protein glycosylation is a well-established post translational modification occurring in all forms of life. In the past two decades, protein glycosylation has been extensively studied in bacterial pathogens underscoring its importance in virulence and colonisation. However, despite the wealth of information regarding protein glycosylation in bacterial pathogens, little is known about this process in gut commensal bacteria. The gut microbiota has co-evolved with and is largely adapted to its host, leading to mutually beneficial interactions. These interactions often involve the adhesion of the bacteria to the gastrointestinal tract using specialised adhesins.
Lactobacillus reuteri is a common gut symbiont found in a wide range of vertebrate hosts. As such, it is used as a model organism to study the co-evolution between gut commensal bacteria and their hosts. Here, we used L. reuteri 100-23 (rodent isolate), ATCC 53608 (pig isolate) and MM4-1a (human isolate) to study the glycosylation of proteins in gut commensal bacteria. An initial bioinformatics approach to identify putative glycosylation clusters suggested the presence of two putative O-glycosylation systems (gtf1/gtf2 and secA2/Y2) that could be involved in post translational modification of proteins in L. reuteri strains. Further genetic and biochemical analyses suggested that Gtf1 is involved in a general glycosylation system targeting multiple proteins in L. reuteri, whereas proteins encoded form the secA2Y2 cluster are dedicated to the glycosylation of a serine rich repeat protein (SRRP). Lectin screening of the secreted proteins from L. reuteri combined with mass spectrometry analysis identified the mucus binding protein MUB53608, SRRP53608 and SRRP100-23, the major adhesins of L. reuteri ATCC 53608 and 100-23, respectively, as glycoproteins. MUB53608 is a putative target of Gtf1-mediated glycosylation. The glycans present on this adhesin were biochemically characterised and found to carry α-galactose and galactofuranose epitopes that could be involved in interactions with the host immune system. The glycosylation of the SRRPs was studied by mass spectrometry and biochemical assays. SRRP53608 harboured GlaNAcα-GlcNAc moieties, whereas SRRP100-23 was found to be glycosylated with Glc-Glc-GlcNAc trisaccharides. The data produced in this work provided novel insights into the L. reuteri glycosylation systems, the nature of glycoproteins and the structure of their glycans, furthering our understanding of the underpinning mechanisms behind their beneficial interactions with the host.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Users 9280 not found.
Date Deposited: 23 Mar 2018 14:27
Last Modified: 28 Feb 2021 01:38


Downloads per month over past year

Actions (login required)

View Item View Item