Proteomic-based analytical approach for the characterization of glutenin subunits in durum wheat
Proteomic-based analytical approach for the characterization of glutenin subunits in durum wheat
复制标题
DOI：
10.1002/jms.1680
复制
发表时间：
2009-12-01
影响因子：
2.3
通讯作者：
Ferranti, Pasquale
中科院分区：
文献类型：
--
作者：
Mamone, Gianfranco;De Caro, Salvatore;Ferranti, Pasquale
小麦谷蛋白亚基分析的主要目的之一是鉴定与小麦品质相关的蛋白质组分。谷蛋白的蛋白质组学特征必须考虑相对低水平的精氨酸和赖氨酸残基以及这些亚基的不同基团之间的紧密序列相似性，这阻碍甚至阻止了GS的鉴定。本研究应用蛋白质组学方法对小麦谷蛋白组分的异质性进行了研究。首次采用双向凝胶电泳技术分析了从硬粒小麦粉中提取的蛋白质，大大降低了谷蛋白的复杂性。通过胰蛋白酶肽的纳米液相色谱串联质谱分析确认每个斑点的身份。同时，通过基质辅助激光解吸/电离飞行时间分析测量高质量范围允许检测大胰蛋白酶肽。收集来自搜索引擎询问的所有数据，获得了高分子量GS（包括Bx7和By 8）的非常高的序列覆盖率，与硬粒小麦的已知遗传谱一致。此外，还发现了一个以前从未检测到的By 8截短型，并以合理的序列覆盖率鉴定了低分子量谷蛋白亚基（LMW-GS）B型，而由于小麦DNA数据库的不完整，LMW-GS C型和D型的鉴定受到了限制。本研究首次对小麦谷蛋白质组进行了全面的分析，为小麦品种的分类提供了可靠的方法。版权所有（C）2009约翰威利父子有限公司
One of the main objectives of wheat glutenin subunit (GS) analysis is the identification of protein components linked to wheat quality. The proteomic characterization of glutenin has to consider the relatively low levels of arginine and lysine residues and the close sequence similarity among the different groups of these subunits, which hinders or even prevents the identification of the GS. In this study, a proteomic approach has been applied to resolve the heterogeneity of wheat glutenin components. Proteins extracted from Triticum durum flour were first analyzed by two-dimensional gel electrophoresis, which greatly reduced glutenin complexity. The identity of each spot was confirmed by nano liquid chromatography tandem mass spectrometry analysis of tryptic peptides. In parallel, measurements of the high mass range by matrix-assisted laser desorption/ionization time-of-flight analysis allowed detection of the large tryptic peptides. Gathering all data from search engine interrogation, very high sequence coverage was obtained for high molecular weight GS, including Bx7 and By8, in agreement with the known genetic profile of durum wheat. In addition, a truncated form of By8, never detected before, was also found. Low molecular weight GS (LMW-GS) B-type was identified with reasonable sequence coverage, while a clear identification of LWM-GS C- and D-type was hindered by the incompleteness of the wheat DNA databases. This study represents the first comprehensive analysis of the glutenin proteome and provides a reliable method for classifying wheat varieties according to their glutenin profile. Copyright (C) 2009 John Wiley & Sons, Ltd.