Abstract Methodology for the quantitative analysis of intact glutenin proteins by ESI-LC-MS was developed with 1 Da mass resolution, constituting the first published application of proteoform profiling to plant biology. Two parent lines and 28 F5 crosses were analyzed in two blocks, 14 months apart. Control sample data were used to align retention times and normalize abundance between sample sets. A total of 4622 observations of 347 distinct proteoforms between 17 899 and 88 744 Da were observed. Proteoform abundances spanned a 1000-fold range and were linear (r2 > 0.990) with dilution. A novel method for the objective elimination of low intensity, noise-dominated data using replicate variability within the dataset is presented. Two abundant PTMs were detected; one known but uncharacterized Bx and Dy high-molecular-weight glutenin subunit (HMWGS) PTM and the other in 24 low molecular weight proteoform pairs. Finally, 16 abundant proteoforms were detected in progeny but not in either parent. This application should increase the statistical power of correlations between gluten complement and functional data and drive the detection of novel PTMs that may indicate differential regulation of the cellular processes related to quality.

中文翻译：

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小麦谷蛋白亚基的定量 LC-MS 完整蛋白质型分析

摘要 通过 ESI-LC-MS 对完整麦谷蛋白进行定量分析的方法学以 1 Da 的质量分辨率开发，构成了蛋白质型分析在植物生物学中的首次公开应用。两个亲本系和 28 个 F5 杂交在两个区块中进行了分析，相隔 14 个月。对照样品数据用于对齐保留时间和标准化样品组之间的丰度。在 17 899 到 88 744 Da 之间共观察到 347 种不同的蛋白质组的 4622 次观察结果。蛋白质型丰度跨越 1000 倍的范围，并且在稀释时呈线性 (r2 > 0.990)。提出了一种使用数据集内的重复变异性客观消除低强度、以噪声为主的数据的新方法。检测到两个丰富的 PTM；一个已知但未表征的 Bx 和 Dy 高分子量谷蛋白亚基 (HMWGS) PTM 和另一个在 24 个低分子量蛋白质组对中。最后，在后代中检测到 16 种丰富的蛋白质型，但在任​​一亲本中均未检测到。此应用程序应增加面筋补充和功能数据之间相关性的统计能力，并推动新 PTM 的检测，这些 PTM 可能表明与质量相关的细胞过程的差异调节。