J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida, Gainesville, Florida 32611, United States.
J Phys Chem B. 2021 Jun 24;125(24):6559-6571. doi: 10.1021/acs.jpcb.1c02083. Epub 2021 Jun 15.
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 transmission electron microscopy, fluorescence microscopy, and thioflavin T spectroscopy, we show that glycosylation at a site external to the fibrillization domain can alter the self-assembly pathway of a synthetic fibrillizing peptide, NSGSGQQKFQFQFEQQ (NQ11). Specifically, an NQ11 variant modified with N-linked -acetylglucosamine, N(GlcNAc)SGSG-Q11 (GQ11), formed β-sheet nanofibers more slowly than NQ11 in deionized water (pH 5.8), which correlated to the tendency of GQ11 to form a combination of short fibrils and nonfibrillar aggregates, whereas NQ11 formed extended nanofibers. Acidic phosphate buffer slowed the rate of GQ11 fibrillization and altered the morphology of the structures formed yet had no effect on NQ11 fibrillization rate or morphology. The buffer ionic strength had no effect on the fibrillization rate of either peptide, while the diphosphate anion had a similar effect on the rate of fibrillization of both peptides. Collectively, these data demonstrate that a glycan moiety located external to the β-sheet fibrillizing domain can alter the pH-dependent 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 effect of glycosylation on peptide self-assembly and should guide future efforts to develop biomaterials from synthetic β-sheet fibrillizing glycopeptides.
由于其生物相容性和可生物降解性，自组装成长纤维的短合成肽（即肽纳米纤维）被广泛用于创建用于各种医学和生物技术应用的生物材料。糖基化是一种常见的蛋白质翻译后修饰，它越来越受到关注，因为它可以创建能够模拟天然碳水化合物修饰蛋白功能的肽纳米纤维。最近的报告表明，糖基化可以破坏天然淀粉样肽的纤维化。在这里，我们使用透射电子显微镜、荧光显微镜和硫黄素 T 光谱法，表明在纤维化结构域之外的部位进行糖基化可以改变合成纤维化肽 NSGSGQQKFQFQFEQQ（NQ11）的自组装途径。具体来说，用 N-连接的 N-乙酰葡萄糖胺修饰的 NQ11 变体，N（GlcNAc）SGSG-Q11（GQ11），在去离子水中（pH5.8）比 NQ11 形成 β-片层纳米纤维的速度更慢，这与 GQ11 形成短纤维和无纤维聚集物的组合的趋势相关，而 NQ11 形成延伸的纳米纤维。酸性磷酸盐缓冲液减缓了 GQ11 纤维化的速度并改变了形成的结构的形态，但对 NQ11 纤维化的速度或形态没有影响。缓冲液离子强度对两种肽的纤维化速度均无影响，而二磷酸阴离子对两种肽的纤维化速度均有类似的影响。总的来说，这些数据表明，位于β-片层纤维化结构域之外的糖基部分可以改变合成肽的 pH 依赖性自组装途径，导致形成的纤维质量和形态发生显著变化。这些观察结果增加了对糖基化对肽自组装影响的理解，并应指导未来从合成β-片层纤维化糖肽开发生物材料的努力。
