Rapid separation and characterization of grain water-soluble proteins in bread wheat cultivars (Triticum aestivum L.) by capillary electrophoresis

Abstract

. 2008. Rapid separation and characterization of grain water-soluble proteins in bread wheat cultivars (Triticum aestivum L.) by capillary electrophoresis. Can. J. Plant Sci. 88: 843Á848. Water-soluble (WS) proteins in wheat grain are considered to represent the suite of biologically active enzymes and enzyme inhibitors in the grain. In this study, a rapid capillary electrophoresis (CE) method for WS protein separations was developed using untreated fused-silica columns and an acidic phosphate-glycine buffer system. In order to optimize the resolution and reproducibility of CE separation, different protein extraction methods, organic modifiers in phosphate-glycine buffer and capillary electrophoresis conditions, including capillary length and inner diameter (ID), operating temperature, performance voltages, sample injection times, etc., were investigated. High resolution and reproducibility of WS proteins were achieved using 20% ethanol as the extracting buffer. The optimal condition to separate these proteins was 50 mm ID)31.5 cm (26.5 cm to the detector) capillary at 11.0 kV and 358C. The optimum buffer was 0.1 M phosphate-glycine (pH 2.5) containing 20% acetonitrile (ACN) and 0.05% hydroxylpropylmethylcellulose. Using this method, the WS proteins were well separated in less than 10 min. A total of 120 Chinese bread wheat cultivars were analyzed. The CE patterns of most bread wheat cultivars showed a higher level of polymorphisms compared with SDS-PAGE patterns. All cultivars analyzed could be readily differentiated based on their WS protein profiles. Results indicate that the WS proteins are useful biochemical markers for wheat genetics and breeding research and CE is expected to become a new and powerful tool for the separation and characterization of grain WS proteins in bread wheat.Key words: Triticum aestivum, bread wheat, water-soluble proteins, capillary electrophoresis, biochemical markers Wang, A., Pei, Y., Li, X., Zhang, Y., Zhang, Q., He, Z., Xia, X., Appels, R., Ma, W., Huang, X.-Q. et Yan, Y. 2008. Se´paration et caracte´risation rapides des prote´ines hydrosolubles dans le grain des cultivars de ble´tendre (Triticum aestivum L.) par e´lectrophore`se capillaire. Can. J. Plant Sci. 88: 843Á848. Les prote´ines hydrosolubles, estime-t-on, correspondent al 'ensemble des enzymes biologiquement actives et des inhibiteurs des enzymes pre´sents dans le grain du ble´tendre. Les auteurs ont mis au point une technique rapide d'e´lectrophore`se capillaire (EC) pour se´parer les prote´ines hydrosolubles en recourant a`des colonnes de silice fondue et a`une solution de phosphate acide tamponne´e a`la glycine. Pour la meilleure re´solution et la reproductibilite´optimale de la se´paration par EC, ils ont examine´diverses me´thodes d'extraction des prote´ines, plusieurs modificateurs organiques du tampon phosphate-glycine et diffe´rents parame`tres de l'e´lectrophore`se capillaire, parmi lesquels la longueur et le diame`tre inte´rieur (DI) des capillaires, la tempe´rature et la tension durant l'ope´ration et la d...

Characterization and Comparative Analysis of Wheat High Molecular Weight Glutenin Subunits by SDS-PAGE, RP-HPLC, HPCE, and MALDI-TOF-MS

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Electrophoretic and electrochromatographic separation of proteins in capillaries: An update covering 2007–2009

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