Location: Crop Improvement & Utilization Research
Title: Analysis of expressed sequence tags from a single wheat cultivar facilitates interpretation of tandem mass spectrometry data and discrimination of gamma gliadin proteins that may play different functional roles in flour Authors
Submitted to: Biomed Central (BMC) Plant Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 11, 2010
Publication Date: January 11, 2010
Citation: Altenbach, S.B., Vensel, W.H., Dupont, F.M. 2010. Analysis of expressed sequence tags from a single wheat cultivar facilitates interpretation of tandem mass spectrometry data and discrimination of gamma gliadin proteins that may play different functional roles in flour. Biomed Central (BMC) Plant Biology. 10(7):1471-2229. Available: http://www.biomedcentral.com/1471-2229/10/7. Interpretive Summary: The gamma gliadins are a complex group of proteins that together with other gluten proteins determine the functional properties of wheat flour. The proteins have unusual amino acid compositions and contain large regions of repetitive sequences. While most gamma gliadins are present in the flour as single proteins, some contain minor sequence variations that enable them to be linked with other gluten proteins into large polymers that are critical for flour quality. The ability to differentiate among the gamma gliadins is important for studies of flour quality because proteins with similar sequences can have different effects on functional properties. Mass spectrometry is often used to identify proteins. However, it is challenging to use mass spectrometry to identify the wheat gamma gliadins because of the complexity of the proteins and the lack of complete sequence information in databases that are used to evaluate mass spectrometry data. To overcome these problems, a detailed analysis of gamma gliadin genes expressed in a US wheat cultivar was performed. The inclusion of these cultivar-specific sequences in databases used to analyze mass spectrometry data made it possible to distinguish individual gamma gliadins that play different roles in flour quality.
Technical Abstract: The complement of gamma gliadin genes expressed in the wheat cultivar Butte 86 was evaluated by analyzing publicly available expressed sequence tag (EST) data. Eleven contigs were assembled from 153 Butte 86 ESTs. Nine of the contigs encoded full-length proteins and four of the proteins contained an extra cysteine residue that would enable them to be linked with other gluten proteins into large polymers critical for flour quality. Only one of the encoded proteins was a perfect match with a sequence reported in NCBI. Contigs from four different publicly available EST assemblies encoded proteins that were perfect matches with some, but not all, of the Butte 86 gamma gliadins and the complement of identical proteins was different for each assembly. A specialized database that included the sequences of Butte 86 gamma gliadins was constructed for identification of flour proteins by tandem mass spectrometry (MS/MS). In a pilot experiment, proteins corresponding to six Butte 86 gamma gliadin contigs were distinguished by MS/MS, including one containing the extra cysteine residue. Two other proteins were identified as one of two closely related Butte 86 proteins but could not be distinguished unequivocally. Unique peptide tags specific for Butte 86 gamma gliadins are reported.