Yen Hsin-Yung, Liko Idlir, Gault Joseph, Wu Di, Struwe Weston B, Robinson Carol V
Physical and Theoretical Chemistry, Department of Chemistry, University of Oxford, South Parks Road, Oxford, OX1 3TA, UK.
Present address: OMass Therapeutics, The Schrodinger Building, Oxford Science Park, Oxford, OX4 4GE, UK.
Angew Chem Int Ed Engl. 2020 Sep 1;59(36):15560-15564. doi: 10.1002/anie.202005727. Epub 2020 Jul 7.
The immune scavenger protein DC-SIGN interacts with glycosylated proteins and has a putative role in facilitating viral infection. How these recognition events take place with different viruses is not clear and the effects of glycosylation on the folding and stability of DC-SIGN have not been reported. Herein, we report the development and application of a mass-spectrometry-based approach to both uncover and characterise the effects of O-glycans on the stability of DC-SIGN. We first quantify the Core 1 and 2 O-glycan structures on the carbohydrate recognition and extracellular domains of the protein using sequential exoglycosidase sequencing. Using ion mobility mass spectrometry, we show how specific O-glycans, and/or single monosaccharide substitutions, alter both the overall collision cross section and the gas-phase stability of the DC-SIGN isoforms. We find that rather than the mass or length of glycoprotein modifications, the stability of DC-SIGN is better correlated with the number of glycosylation sites.
免疫清除蛋白DC-SIGN与糖基化蛋白相互作用，在促进病毒感染方面可能发挥作用。目前尚不清楚这些识别事件如何在不同病毒中发生，且糖基化对DC-SIGN折叠和稳定性的影响尚未见报道。在此，我们报告了一种基于质谱的方法的开发与应用，用于揭示和表征O-聚糖对DC-SIGN稳定性的影响。我们首先使用外切糖苷酶序列分析对该蛋白碳水化合物识别和细胞外结构域上的核心1和核心2 O-聚糖结构进行定量。通过离子淌度质谱，我们展示了特定的O-聚糖和/或单糖取代如何改变DC-SIGN异构体的整体碰撞截面和气相稳定性。我们发现，DC-SIGN的稳定性与糖基化位点的数量相关性更好，而非糖蛋白修饰的质量或长度。
