色譜保留時(shí)間在蛋白質(zhì)組研究中的應(yīng)用

邵 晨, 高友鶴

(中國(guó)醫(yī)學(xué)科學(xué)院基礎(chǔ)醫(yī)學(xué)研究所,中國(guó)協(xié)和醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院,生理和病理生理學(xué)系,北京100005)

色譜保留時(shí)間在蛋白質(zhì)組研究中的應(yīng)用

邵 晨＊, 高友鶴

(中國(guó)醫(yī)學(xué)科學(xué)院基礎(chǔ)醫(yī)學(xué)研究所,中國(guó)協(xié)和醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院,生理和病理生理學(xué)系,北京100005)

液相色譜與串聯(lián)質(zhì)譜聯(lián)用(LC-MS/MS)技術(shù)是蛋白質(zhì)組學(xué)研究中的常見(jiàn)方法。保留時(shí)間作為獨(dú)立于質(zhì)譜信息的參數(shù)已經(jīng)被用于蛋白質(zhì)的鑒定和定量工作中。在多肽鑒定領(lǐng)域,多肽的色譜保留時(shí)間預(yù)測(cè)與常規(guī)的二級(jí)串聯(lián)質(zhì)譜數(shù)據(jù)庫(kù)搜索算法結(jié)合可以提高鑒定的可信度。鑒定的靈敏度也可以通過(guò)匹配多次LC-MS實(shí)驗(yàn)中具有相同精確質(zhì)量數(shù)和保留時(shí)間的峰而提高。另一方面,由于色譜條件的微小改變即會(huì)引起保留時(shí)間的變化,因此對(duì)多次實(shí)驗(yàn)結(jié)果進(jìn)行保留時(shí)間比對(duì)是進(jìn)行非標(biāo)記定量的不可或缺的步驟。另外,聯(lián)合保留時(shí)間偏移和質(zhì)量數(shù)信息還可以進(jìn)行蛋白質(zhì)翻譯后修飾(post-translational modification,PTM)的鑒定。

液相色譜-串聯(lián)質(zhì)譜;保留時(shí)間預(yù)測(cè);保留時(shí)間比對(duì);多肽鑒定;非標(biāo)記定量;翻譯后修飾;蛋白質(zhì)組學(xué)

Abstract:Liquid Chromatography coup led with tandem mass spectrometry(LC-MS/MS)has been one of the most popular approaches inproteome analysis.As an independent parameter to mass spectrometry information,peptide retention time has been utilized to facilitate protein identification and quantification.In the field of pep tide identification,the prediction of the retention time combined with routine tandem mass spectrometry database searching methods could help improve the confidence of identification.The sensitivity of identification could also be improved by matching peaks with both the accurate mass and retention time in multiple aligned LC-MS runs.Meanwhile,because small changes of liquid Chromatography conditions lead to variability in retention times unavoidably,retention time alignm ent is crucial to label free quantification.Additionally,post-translational modifications(PTM)could be identified by com bining retention time shifts and mass deviation information.

Key words:liquid Chromatography-tandem mass spectrometry(LC-MS/MS);retention time prediction;retention time alignment;peptideidentification;label-free quantification;posttranslational modification(PTM);proteomics

在蛋白質(zhì)組的研究中,液相色譜串聯(lián)一級(jí)質(zhì)譜(liquid Chromatography coupled with mass spectrom etry,LC-MS)或二級(jí)質(zhì)譜(liquid Chromatography coup led with tandem mass spectrometry,LC-MS/MS)是進(jìn)行蛋白質(zhì)鑒定和定量分析的常見(jiàn)策略。其主要實(shí)驗(yàn)流程為:首先將蛋白質(zhì)混合物經(jīng)酶切變?yōu)槎嚯幕旌衔?然后通過(guò)一維或二維色譜分離多肽混合物,再用質(zhì)譜儀鑒定其序列及進(jìn)行定量分析。用一維色譜分離多肽混合物時(shí),通常是利用反……