化学
转铁蛋白
细胞毒性
糖蛋白
唾液酸
质谱法
离子迁移光谱法
生物化学
转铁蛋白受体
凝胶电泳
生物物理学
色谱法
体外
生物
作者
Gongyu Li,Ashley Phetsanthad,M Ma,Qinying Yu,Ashita Nair,Zhen Zheng,Fengfei Ma,Kellen DeLaney,Seungpyo Hong,Lingjun Li
标识
DOI:10.1021/acs.analchem.1c04503
摘要
Protein sialylation has been closely linked to many diseases including Alzheimer's disease (AD). It is also broadly implicated in therapeutics operating in a pattern-dependent (e.g., Neu5Ac vs Neu5Gc) manner. However, how the sialylation pattern affects the AD-associated, transferrin-assisted iron/Aβ cellular uptake process remains largely ill-defined. Herein, we report the use of native ion mobility–mass spectrometry (IM–MS)-based fast structural probing methodology, enabling well-controlled, synergistic, and in situ manipulation of mature glycoproteins and attached sialic acids. IM–MS-centered experiments enable the combinatorial interrogation of sialylation effects on Aβ cytotoxicity and the chemical, conformational, and topological stabilities of transferrin. Cell viability experiments suggest that Neu5Gc replacement enhances the transferrin-assisted, iron loading-associated Aβ cytotoxicity. Native gel electrophoresis and IM–MS reveal that sialylation stabilizes transferrin conformation but inhibits its dimerization. Collectively, IM–MS is adapted to capture key sialylation intermediates involved in fine-tuning AD-associated glycoprotein structural microheterogeneity. Our results provide the molecular basis for the importance of sustaining moderate TF sialylation levels, especially Neu5Ac, in promoting iron cellular transportation and rescuing iron-enhanced Aβ cytotoxicity.
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