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An eco-friendly, low-cost, and automated strategy for phosphoproteome profiling

文献类型： 外文期刊

作者： Zhang, Wenyang 1 ; Lai, Cheuk-Kuen 2 ; Huang, Wenjie 1 ; Li, Wenyan 1 ; Wu, Shaowen 1 ; Kong, Qian 1 ; Hopkinson, Alan C. 2 ; Fernie, Alisdair R. 4 ; Siu, K. W. Michael 2 ; Yan, Shijuan 1 ;

作者机构： 1.Guangdong Acad Agr Sci, Agrobiol Gene Res Ctr, Guangdong Key Lab Crop Germplasm Resources Preserv, Guangzhou 510640, Peoples R China

2.York Univ, Dept Chem, Toronto, ON M3J 1P3, Canada

3.York Univ, Ctr Res Mass Spectrometry, Toronto, ON M3J 1P3, Canada

4.Max Planck Inst Mol Plant Physiol, Muhlenberg 1, D-14476 Potsdam, Germany

5.Univ Windsor, Dept Chem & Biochem, Windsor, ON N9B 3P4, Canada

期刊名称：GREEN CHEMISTRY （ 影响因子：11.034； 五年影响因子：10.794 ）

ISSN： 1463-9262

年卷期： 2022 年 24 卷 24 期

页码：

收录情况： SCI

摘要： Reversible protein phosphorylation plays a vital role in many biological processes. In current phosphoproteomics, targeted enrichment before mass spectrometry (MS) analysis is essential. Many kinds of materials have been developed for phosphopeptide enrichment, but these typically involve complex fabrication process and high consumption of chemicals. Furthermore, the enrichment of phosphopeptides is offline and requires multiple manual steps, thereby making it a tedious workflow with high consumption of organic solvents and plastic consumables. To alleviate these problems, we propose a set of eco-friendly strategies for phosphoproteome profiling by developing a reusable phos-trap column via a one-pot mild sol-gel process and establishing an automated online analytical platform with low cost and low waste. This platform also demonstrated high phosphosite coverage, sensitivity, selectivity, and a wide linear range when a standard phosphoprotein (alpha-casein) was used for the initial development assessment. In subsequent analyses of HeLa cells and maize seedlings, a total of 6575 and 3741 phosphorylation sites were characterized, respectively, using three single-shot technical replicates for each sample type with excellent qualitative and quantitative reproducibility. Taken together, this phos-trap column and automated online analytical platform are expected to bring environmental and low-cost technological advancements to phosphoproteomics, especially for large-scale tasks.

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