Complementary multiple hydrogen-bond-based magnetic composite microspheres for high coverage and efficient phosphopeptide enrichment in bio-samples.
Complementary multiple hydrogen-bond-based magnetic composite microspheres for high coverage and efficient phosphopeptide enrichment in bio-samples.
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DOI：
10.1039/d0tb01410a
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发表时间：
2020-09
期刊：
影响因子：
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通讯作者：
Bin Luo;Lingzhu Yu;Zhiyu Li;Jia He;Chunjie Li;Fang Lan;Yao Wu
中科院分区：
文献类型：
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作者：
Bin Luo;Lingzhu Yu;Zhiyu Li;Jia He;Chunjie Li;Fang Lan;Yao Wu
由于磷酸化位点的数量，单磷酸肽和多磷酸肽表现出显著不同的生物学效应。因此，单磷酸肽和多磷酸肽的综合概况对于分析这些生物和病理过程至关重要。然而，最常用的基于金属氧化物亲和色谱（MOAC）的亲和材料对单磷酸肽表现出更强的选择性，从而丢失了大部分关于多磷酸肽的信息。在这里，我们报告聚合物功能化的磁性纳米复合微球作为一个理想的平台，有效地丰富单和多磷酸肽从复杂的生物样品。在互补的多重氢键相互作用的驱动下，复合微球对模型蛋白和真实的生物样品中的磷酸肽具有显著的富集性能。该方法具有良好的选择性（非磷酸肽与磷酸肽的摩尔比为5000：1）、高富集灵敏度（2 fmol）和高覆盖率，以及对多磷酸肽的高捕获率，显示了其在磷酸化蛋白质组学研究中的巨大潜力。更重要的是，我们成功地捕获了癌症相关的磷酸肽（来自磷蛋白Stathmin-1），并从口腔癌患者的唾液和组织裂解物中鉴定了其相关的磷酸化位点，证明了这种材料用于磷酸化疾病标志物检测和发现的潜力。
Due to the number of phosphorylation sites, mono- and multiple-phosphopeptides exhibit significantly different biological effects. Therefore, comprehensive profiles of mono- and multiple-phosphopeptides are vital for the analysis of these biological and pathological processes. However, the most commonly used affinity materials based on metal oxide affinity chromatography (MOAC) show stronger selectivity toward mono-phosphopeptides, thus losing most information on multiple-phosphopeptides. Herein, we report polymer functionalized magnetic nanocomposite microspheres as an ideal platform to efficiently enrich both mono- and multiple-phosphopeptides from complex biological samples. Driven by complementary multiple hydrogen bonding interactions, the composite microspheres exhibited remarkable performance for phosphopeptide enrichment from model proteins and real bio-samples. Excellent selectivity (the molar ratio of nonphosphopeptides/phosphopeptides was 5000 : 1), high enrichment sensitivity (2 fmol) and coverage, as well as high capture rates of multiple-phosphopeptides revealed their great potential in comprehensive phosphoproteomics studies. More importantly, we successfully captured the cancer related phosphopeptides (from the phosphoprotein Stathmin-1) and identified their relevant phosphorylation sites from oral carcinoma patients' saliva and tissue lysate, demonstrating the potential of this material for phosphorylated disease marker detection and discovery.