Delft University of Technology, Department of Biotechnology, Delft, The Netherlands.
SciBear OU, Tallinn, Estonia.
ISME J. 2022 Feb;16(2):346-357. doi: 10.1038/s41396-021-01073-y. Epub 2021 Aug 2.
The enormous chemical diversity and strain variability of prokaryotic protein glycosylation makes their large-scale exploration exceptionally challenging. Therefore, despite the universal relevance of protein glycosylation across all domains of life, the understanding of their biological significance and the evolutionary forces shaping oligosaccharide structures remains highly limited. Here, we report on a newly established mass binning glycoproteomics approach that establishes the chemical identity of the carbohydrate components and performs untargeted exploration of prokaryotic oligosaccharides from large-scale proteomics data directly. We demonstrate our approach by exploring an enrichment culture of the globally relevant anaerobic ammonium-oxidizing bacterium Ca. Kuenenia stuttgartiensis. By doing so we resolve a remarkable array of oligosaccharides, which are produced by two seemingly unrelated biosynthetic routes, and which modify the same surface-layer protein simultaneously. More intriguingly, the investigated strain also accomplished modulation of highly specialized sugars, supposedly in response to its energy metabolism-the anaerobic oxidation of ammonium-which depends on the acquisition of substrates of opposite charges. Ultimately, we provide a systematic approach for the compositional exploration of prokaryotic protein glycosylation, and reveal a remarkable example for the evolution of complex oligosaccharides in bacteria.
原核生物蛋白糖基化具有巨大的化学多样性和菌株可变性，这使得大规模探索变得异常具有挑战性。因此，尽管蛋白质糖基化在所有生命领域都具有普遍意义，但对其生物学意义和塑造寡糖结构的进化力量的理解仍然非常有限。在这里，我们报告了一种新建立的质量-bin 糖组学方法，该方法可以确定碳水化合物成分的化学特性，并直接从大规模蛋白质组学数据中对原核寡糖进行非靶向探索。我们通过探索全球相关的厌氧氨氧化菌 Ca. Kuenenia stuttgartiensis 的富集培养来证明我们的方法。通过这样做，我们解析了一系列令人瞩目的寡糖，这些寡糖由两种看似不相关的生物合成途径产生，并同时修饰同一种表面层蛋白。更有趣的是，所研究的菌株还能够调节高度专门的糖，这可能是对其能量代谢的反应——即铵的厌氧氧化，这取决于获取相反电荷的底物。最终，我们为原核生物蛋白糖基化的组成探索提供了一种系统的方法，并揭示了细菌中复杂寡糖进化的一个显著例子。
