Deciphering the Roles of Glycosylation in Prokaryotic Biofilm Formation
Biochemistry
Final Report Abstract
Post-translational modifications (PTMs) increase the complexity of an organism’s proteome by several orders of magnitude and are involved in a multitude of protein functions and regulatory mechanisms. Despite their importance, our understanding of PTMs in prokaryotes is very limited. This is especially true for protein glycosylation, a highly complex modification that exhibits vast variability across prokaryotes and has been shown to be involved in the virulence of several pathogenic bacteria. In this project, we aimed for the large-scale, mass spectrometric identification of prokaryotic glycoproteins and the functional analysis of individual glycoproteins. We focused especially on the roles of glycoproteins in the formation of biofilms, microbial communities surrounded by extracellular polymeric matrixes, which are of great medical, economic and ecological significance. While the quantitative, in detail characterization of biofilm formation is still ongoing, we have analyzed the glycoproteome of the model archaeon Haloferax volcanii. With 188 glycopeptides, corresponding to 48 glycoproteins, we identified the as of yet largest glycoproteome in archaea. In this process, a surprising complexity of glycosylation in H. volcanii was revealed, which poses new questions about the regulation of different glycosylation pathways in this organism. Following up on the identification of the largest protein of H. volcanii as a glycoprotein, we showed that this protein is involved in biofilm formation. In addition, a 3D-printed device was developed that will simplify the analysis of biofilm formation by facilitating to monitor cultures while also taking samples of the biofilm at different time points. The simple, versatile, and costefficient nature of this device will allow for its use in a variety of experimental setups. Furthermore, we initiated the Archaeal Proteome Project (ArcPP), a collaborative effort aimed at the comprehensive analysis of proteomics datasets within the archaeal community. This combined analysis not only allowed for the identification of 72% of the H. volcanii proteome but also provided new insights into the cell biology of this haloarchaeon. Through the straightforward extendibility of ArcPP new datasets can be readily integrated and compared to existing datasets in its online database (https://archaealproteomeproject.org/) which provides an invaluable resource for archaeal community. Finally, we developed bioinformatic tools that allow for the universal, discovery-driven identification of glycoproteins. This is of special importance in prokaryotes, since a lot of their glycosylation pathways, and consequentially their glycan compositions, are unknown. Facilitating their analysis sets the stage for a better understanding of their functions.
Publications
- (2020) The Archaeal Proteome Project advances knowledge about archaeal cell biology through comprehensive proteomics. Nature communications 11 (1) 3145
Schulze, Stefan; Adams, Zachary; Cerletti, Micaela; Castro, Rosana de; Ferreira-Cerca, Sébastien; Fufezan, Christian; Giménez, María Inés; Hippler, Michael; Jevtic, Zivojin; Knüppel, Robert; Legerme, Georgio; Lenz, Christof; Marchfelder, Anita; Maupin-Fur
(See online at https://doi.org/10.1038/s41467-020-16784-7) - (2018). Archaeal cell surface biogenesis. FEMS Microbiol Rev 42(5)
M. Pohlschroder, F. Pfeiffer, S. Schulze, M.F. Abdul Halim
(See online at https://doi.org/10.1093/femsre/fuy027) - (2019). Haloferax volcanii. Trends Microbiol 27(1)
M. Pohlschroder, S. Schulze
(See online at https://doi.org/10.1016/j.tim.2018.10.004) - (2020). Lipid Anchoring of Archaeosortase Substrates and Midcell Growth in Haloarchaea. mBio 11(2)
M. F. Abdul-Halim, S. Schulze, A. DiLucido, F. Pfeiffer, A. W. Bisson Filho, M. Pohlschroder
(See online at https://doi.org/10.1128/mBio.00349-20)
