Owing to their biocompatibility and biodegradability, short synthetic peptides that self-assemble into elongated β-sheet fibers (i.e., peptide nanofibers) are widely used to create biomaterials for diverse medical and biotechnology applications. Glycosylation, which is a common protein post-translational modification, is gaining interest for creating peptide nanofibers that can mimic the function of natural carbohydrate-modified proteins. Recent reports have shown that glycosylation can disrupt the fibrillization of natural amyloid-forming peptides. Here, using a synthetic fibrillizing peptide, NSGSGQQKFQFQFEQQ (NQ11), we show that glycosylation at a site external to the fibrillization domain can alter the pathway of peptide nanofiber formation. Using transmission electron microscopy, fluorescence microscopy, and thioflavin T spectroscopy, we show that an NQ11 variant modified with N-linked N-acetylglucosamine, N(GlcNAc)SGSG-Q11 (GQ11), formed β-sheet nanofibers more slowly than NQ11 in deionized water (pH 5.8) and acidic phosphate buffer (pH 3.6 and 5.8). Buffer ionic strength had no effect on fibrillization rate, while diphosphate anion had a similar effect on the rate of fibrillization of both GQ11 and NQ11. In deionized water, GQ11 formed a combination of short fibrils and non-fibrillar aggregates, whereas NQ11 formed long nanofibers. At pH 3.6 in phosphate buffer, GQ11 formed short fibrils whereas NQ11 formed long fibers and mats. Collectively, these data demonstrate that a glycan moiety located external to the b-sheet fibrillizing domain can alter the self-assembly pathway of a synthetic peptide, leading to significant changes in the fibril mass and morphology of the structures formed. These observations add to the understanding of the effects of glycosylation on peptide self-assembly, and should guide future efforts to develop biomaterials from synthetic β-sheet fibrillizing glycopeptides.

Glycosylation of a non-fibrillizing appendage alters the self-assembly pathway of a synthetic β-sheet fibrillizing peptide

ACS Fall 2021

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