Selective enrichment of endogenous peptides by chemically modified porous nanoparticles for peptidome analysis.
Selective enrichment of endogenous peptides by chemically modified porous nanoparticles for peptidome analysis.
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DOI：
10.1016/j.chroma.2008.10.002
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发表时间：
2009-02
期刊：
影响因子：
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通讯作者：
Ruijun Tian;Lianbing Ren;Huaijun Ma;Xin Li;Lianghai Hu;M. Ye;R. Wu;Zhijian Tian;Zhen Liu
中科院分区：
文献类型：
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作者：
Ruijun Tian;Lianbing Ren;Huaijun Ma;Xin Li;Lianghai Hu;M. Ye;R. Wu;Zhijian Tian;Zhen Liu
我们报告了一种在自然pH条件下从复杂生物样品中高容量、全面富集内源性肽的组合策略的发展。合成了具有高度有序的纳米孔（即4.8nm）和高比表面积（即751 m2/g）的MCM-41纳米粒子，并用强阳离子交换（SCX-MCM-41）和强阴离子交换（SAX-MCM-41）基团对其进行了修饰。修饰后的纳米粒子对分子量约为100的标准蛋白溶菌酶具有良好的尺寸排阻效应。分子量小于15 kDa的肽段能被很好地吸附。对5种盐浓度的富集肽进行分步洗脱，结果表明，两种修饰纳米粒子均具有较高的富集能力和互补性，且SAX-MCM-41纳米粒子对等电点（pI）小于4的酸性肽具有明显的高选择性。通过将SCX-MCM-41和SAX-MCM-41与未修饰的MCM-41纳米颗粒进一步组合，实现了2 mg小鼠肝脏提取物中内源性肽的大规模富集。应用在线2D nano-LC/MS/MS对富集样品进行分析，总共鉴定了2721个独特的肽。MW与pI分布的二维分析结合所鉴定的肽的丰度表明，这三种类型的纳米颗粒具有全面的互补性肽组富集。
We report the development of a combined strategy for high capacity, comprehensive enrichment of endogenous peptide from complex biological samples at natural pH condition. MCM-41 nanoparticles with highly ordered nanoscale pores (i.e. 4.8nm) and high-surface area (i.e. 751m2/g) were synthesized and modified with strong cation-exchange (SCX-MCM-41) and strong anion-exchange (SAX-MCM-41) groups. The modified nanoparticles demonstrated good size-exclusion effect for the adsorption of standard protein lysozyme with molecular weight (MW) of ca. 15kDa; and the peptides with MW lower than this value can be well adsorbed. Step elution of the enriched peptides with five salt concentrations presented that both modified nanoparticles have high capacity and complementarity for peptides enrichment, and the SAX-MCM-41 nanoparticles has obviously high selectivity for acidic peptides with pI (isoelectric point) lower than 4. Large-scale enrichment of endogenous peptides in 2mg mouse liver extract was achieved by further combination of SCX-MCM-41 and SAX-MCM-41 with unmodified MCM-41 nanoparticles. On-line 2D nano-LC/MS/MS was applied to analyze the enriched samples, and 2721 unique peptides were identified in total. Two-dimensional analysis of MW versus pI distribution combined with abundance of the identified peptides demonstrated that the three types of nanoparticles have comprehensive complementarity for peptidome enrichment.