Impact of peptide modifications on the isobaric tags for relative and absolute quantitation method accuracy.
Impact of peptide modifications on the isobaric tags for relative and absolute quantitation method accuracy.
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DOI：
10.1021/ac100775s
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发表时间：
2011-02-01
影响因子：
7.4
通讯作者：
Lazar, Iulia M.
中科院分区：
文献类型：
--
作者：
Tenga, Milagros J.;Lazar, Iulia M.
在这项研究中，评估了氨基酸修饰对iTRAQ(相对和绝对定量的等压标签)方法的准确性的影响。以17-雌二醇(17β-estadiol，E_2)和三苯氧胺(Tamoxifen，)联合培养的MCF-7乳腺癌细胞为模型。采用反相高效液相色谱(RPLC)和脉冲Q解离(PQD)离子捕获串联质谱仪(MS/MS)对细胞进行标记和分析。通过使用氨基酸修饰的各种组合，即Lys/Tyr/Cys和氨基末端iTRAQ标记、Lys甲基化、乙酰化和氨甲酰化以及Cys/Met氧化来进行数据库搜索。除了预期的赖氨酸/氨基末端iTRAQ标记，这种修饰是酶或样品制备相关反应的结果，通常在定量分析中被忽略，以最大限度地减少假阳性多肽鉴定的比率。研究表明，对蛋白质鉴定和定量影响最大的修改与赖氨酸和酪氨酸氨基酸残基有关，通过这种修改，识别的蛋白质的数量和类型将发生变化(最多10%)，如果使用适当的数据过滤条件，蛋白质识别的假阳性率可以保持在5%的上限以下。此外，还检查了可能的翻译后修饰(即磷酸化)对iTRAQ定量的干扰。
In this study, the impact of amino acid modifications on the accuracy of the iTRAQ (isobaric tags for relative and absolute quantitation) method was evaluated. MCF-7 breast cancer cells, cultured in the presence of 17 β-estradiol (E2) and tamoxifen (Tam), were used as a model system. The cells were labeled and analyzed by reversed-phase liquid chromatography (RPLC) and pulsed Q dissociation (PQD) ion trap tandem mass spectrometry (MS/MS) detection. Database searching was performed by using various combinations of amino acid modification allowances, i.e, Lys/Tyr/Cys and amino terminal iTRAQ labeling, Lys methylation, acetylation and carbamylation, and Cys/Met oxidation. Other than the intended Lys/amino terminal iTRAQ labeling, such modifications occur as a result of either enzymatic or sample prep related reactions, and are typically ignored in quantitation analysis to minimize the rate of false positive peptide identifications. The study revealed that the modifications with the greatest impact on protein identification and quantitation pertain to Lys and Tyr amino acid residues, that by enabling such modifications the number and type of identified proteins will change (by up to 10 %), and that the rate of false positive protein identifications can be maintained below an upper threshold of 5 % if appropriate data filtering conditions are used. In addition, the interference of possible posttranslational modifications (i.e., phosphorylation) with iTRAQ quantitation was examined.
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影响因子：
2
作者：
Sarvaiya, Hetal A.;Yoon, Jung H.;Lazar, Iulia M.
通讯作者：
Lazar, Iulia M.
影响因子：
7
作者：
Ross, PL;Huang, YLN;Pappin, DJ
通讯作者：
Pappin, DJ
DOI：
10.1016/j.biocel.2007.03.007
发表时间：
2007-01-01
影响因子：
4
作者：
Rosati, Alessandra;Ammirante, Massimo;Turco, Maria Caterina
通讯作者：
Turco, Maria Caterina
影响因子：
4.4
作者：
DeSouza, L;Diehl, G;Siu, KWM
通讯作者：
Siu, KWM
DOI：
10.1016/j.jasms.2009.02.029
发表时间：
2009-07-01
影响因子：
3.2
作者：
Armenta, Jenny M.;Hoeschele, Ina;Lazar, Iulia M.
通讯作者：
Lazar, Iulia M.