ChELSI Institute, Biological and Environmental Systems Group, Department of Chemical and Process Engineering, The University of Sheffield, Mappin Street, Sheffield, S1 3JD, United Kingdom.
J Proteome Res. 2010 Feb 5;9(2):1165-72. doi: 10.1021/pr9007688.
A quantitative proteomic analysis of the membrane of the archaeon Sulfolobus solfataricus P2 using iTRAQ was successfully demonstrated in this technical note. The estimated number of membrane proteins of this organism is 883 (predicted based on Gravy score), corresponding to 30% of the total number of proteins. Using a modified iTRAQ protocol for membrane protein analysis, of the 284 proteins detected, 246 proteins were identified as membrane proteins, while using an original iTRAQ protocol, 147 proteins were detected with only 133 proteins being identified as membrane proteins. Furthermore, 97.2% of proteins identified in the modified protocol contained more than 2 distinct peptides compared to the original workflow. The successful application of this modified protocol offers a potential technique for quantitatively analyzing membrane-associated proteomes of organisms in the archaeal kingdom. The combination of 3 different iTRAQ experiments resulted in the detection of 395 proteins (>or=2 distinct peptides) of which 373 had predicted membrane properties. Approximately 20% of the quantified proteins were observed to exhibit >or=1.5-fold differential expression at temperatures well below the optimum for growth.
本技术说明成功展示了使用 iTRAQ 对古菌 Sulfolobus solfataricus P2 的膜进行定量蛋白质组分析。该生物的膜蛋白估计数量为 883 个（根据 Gravy 评分预测），占总蛋白数的 30%。使用改良的 iTRAQ 膜蛋白分析方案，在检测到的 284 种蛋白质中，有 246 种被鉴定为膜蛋白，而使用原始的 iTRAQ 方案，仅检测到 147 种蛋白质，其中只有 133 种被鉴定为膜蛋白。此外，与原始工作流程相比，改良方案中鉴定的 97.2%的蛋白质包含超过 2 个不同的肽段。该改良方案的成功应用为定量分析古菌王国中与膜相关的蛋白质组提供了一种潜在技术。3 种不同 iTRAQ 实验的组合检测到了 395 种（≥2 个不同肽段）蛋白质，其中 373 种具有预测的膜特性。约 20%的定量蛋白质在远低于生长最适温度下表现出≥1.5 倍的差异表达。
