Jun 28, 2022, 4:50 PM
Room: S1

Room: S1


Brito, Alexandra (3B’s Research Group, I3Bs - Research Institute on Biomaterials, Biodegradables and Biomimetics, University of Minho )


"Introduction: Folding is a crucial process that modulates the function of proteins:[1] non-covalent intramolecular interactions between amino acids give rise to a defined three-dimensional structure with minimal free energy known as protein native state. This is an error prompt process that often results in misfolding and the formation of off-pathway aggregates associated with pathological conditions. There are different cellular mechanisms to control this process, e.g., post-translational modifications that change the free energy of the protein. An example is glycosylation that affects the protein folding and aggregation.[2] The thermodynamics and kinetics of these processes are still poorly understood and often rely only on in silico models. Herein, we show that O-glycotripeptides are extremely useful reductionist models to study the involvement of glycosylation in these processes at molecular and microscopic levels.[3]

Methodology: Minimalistic glycoproteins were designed using glycosylated serine (S) or threonine (T) flanked by phenylalanine (F). The glycosylated S and T are characteristic structural components of O-glycoproteins, while the aromatic F was introduced to augment the aggregation propensity of the glycopeptides. The aggregation of these glycotripeptides was compared to their respective non-glycosylated analogues using in silico all-atom molecular dynamics simulations and in vitro by circular dichroism (CD) and X-ray diffraction (XRD). The morphology of the generated aggregates was visualized by scanning (SEM) and transmission (TEM) electron microscopies and their mechanical properties were measured by atomic force microscopy (AFM).

Results: We were able to assess the distinct contributions of F, S or T and glucose to the glycopeptides’ stereochemistry and aggregation. Although S and T differ only by a methyl group, this subtle variation affects the inter- and intramolecular CH-pi interactions between F and S or T: F/S << F/T. S to T substitution also induced alterations in the morphology of the generated supramolecular aggregates as shown for the non-glycosylated peptides. O-glycosylation introduced changes in the pi-interactome by establishing additional CH-pi interactions, i.e., Glc/F. The aggregates of the glycopeptides have reduced stiffness and increased thermal stability when compared to their non-glycosylated counterparts. These changes were more prominent for the S analogues when compared with the T ones.

Conclusions: We demonstrate that simple glycotripeptides are a useful model for revealing the mechanism(s) of the aggregation processes at the molecular level. The generated assemblies can be also used as functional biomaterials acting as biomimetics of glycoproteins.

Acknowledgements: We acknowledge the EU's H2020 program (Forecast 668983) and the Portuguese FCT (BD/113794/2015; PTDC/BTM-MAT/28327/2017 CARDIOHEAL; CEECINST/00077/2018) for the financial support. Part of this research was supported by National Science Foundation (NSF) grant CHE-1808143 and a grant of computer time from the City University of New York High Performance Computing Center under NSF grants CNS-0855217, CNS-0958379 and ACI-1126113.

[1] Shental-Bechor, D. et al. PNAS. 24, 8256-8261 (2008).
[2] Xu, C. et al. Nat. Rev. Mol. 16, 742-752 (2015).
[3] Brito, A.B. et al. JACS, 143, 19703-19710 (2021)."


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