The use of accurate mass tags for high-throughput microbial proteomics.
The use of accurate mass tags for high-throughput microbial proteomics.
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使用精确质量标签进行高通量微生物蛋白质组学。
DOI：
10.1089/15362310252780843
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发表时间：
2002
期刊：
影响因子：
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通讯作者：
Udseth,HaroldR
中科院分区：
文献类型：
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作者：
Smith,RichardD;Anderson,GordonA;Lipton,MaryS;Masselon,Christophe;Pasa-Tolic,Ljiljana;Shen,Yufeng;Udseth,HaroldR
我们描述和审查进展的全球战略，旨在扩大灵敏度，动态范围，全面性和通量的蛋白质组学测量的基础上微生物系统 使用由全局蛋白酶消化产生的多肽精确质量标签（AMT）。两阶段的策略利用傅里叶变换离子回旋共振的高精度质量测量 使用串联质谱法（MS/MS）从暂时鉴定的潜在质量标签中验证特定生物体的多肽AMT，为随后的 一个高分辨率的毛细管液相色谱分离结合高灵敏度，高分辨率的准确FTICR测量显示， 能够表征超过105种组分的多肽混合物，足以使用AMT进行广泛的蛋白质鉴定。该方法的优点包括： 蛋白质鉴定，其广泛的蛋白质组覆盖范围，以及稳定同位素标记方法用于精确相对蛋白质丰度测量的能力。对该战略进行了初步评估 使用的是酵母菌和耐辐射球菌。其他发展，包括使用多路MS/MS能力和动态范围扩展方法 的蛋白质组测量的承诺，以进一步扩展蛋白质组学测量的质量，也被描述。
We describe and review progress towards a global strategy that aims to extend the sensitivity, dynamic range, comprehensiveness, and throughput of proteomic measurements for microbial systems based upon the use of polypeptide accurate mass tags (AMTs) produced by global protein enzymatic digestions. The two-stage strategy exploits high accuracy mass measurements using Fourier transform ion cyclotron resonance mass spectrometry (FTICR) to validate polypeptide AMTs for a specific organism, from potential mass tags tentatively identified using tandem mass spectrometry (MS/MS), providing the basis for subsequent measurements without the need for routine MS/MS. A high-resolution capillary liquid chromatography separation combined with high sensitivity, and high-resolution accurate FTICR measurements is shown to be capable of characterizing polypeptide mixtures of more than 105components, sufficient for broad protein identification using AMTs. Advantages of the approach include the high confidence of protein identification, its broad proteome coverage, and the capability for stable-isotope labeling methods for precise relative protein abundance measurements. The strategy has been initially evaluated using the microorganismsSaccharomyces cerevisiaeandDeinococcus radiodurans. Additional developments, including the use of multiplexed-MS/MS capabilities and methods for dynamic range expansion of proteome measurements that promise to further extend the quality of proteomics measurements, are also described.