Characterization of wheat gluten proteins by hplc and MALDI TOF mass spectrometry |
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Authors: | Yuwei Qian Ken Preston Oleg Krokhin Jean Mellish Werner Ens |
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Institution: | (1) Department of Molecular Biology and Genetic Engineering, College of Basic Sciences and Humanities, G. B. Pant University of Agriculture & Technology, Pantnagar, 263 145, India;(2) Molecular Biology & Biotechnology, Central Salt and Marine Chemicals Research Institute, G. B. Marg, Bhavnagar, Gujarat, 364 002, India; |
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Abstract: | We have performed a detailed characterization and identification of wheat gluten proteins obtained from the Teal variety of
Canadian hard red spring wheat. RP-HPLC separation of the sample into 35 fractions has reduced the spectral complexity; this
was followed by MALDI mass spectrometry (MS), which showed the presence of six or fewer resolved protein components above
20 kDa in each RP-HPLC fraction, giving a total of 93 MS resolved peaks. These included 17 peaks in the ω-gliadin fractions (F1–4), 12 in the high molecular weight (HMW) glutenin subunit fractions (F5–8), 59 in the α- and β-gliadins and low molecular weight (LMW) glutenin subunit fractions (F9–31) and 5 peaks in the γ-gliadin fractions (F32–35). Peptide maps of tryptic digests of HPLC fractions were obtained from a tandem quadrupole time-of-flight
mass spectrometer (MALDI QqTOF MS) and were submitted to the ProFound search engine. HMW glutenin subunits including Ax2*,
Dx5, Bx7, and Dy10 (consistent with the known profile of Teal), and LMW glutenin subunits including six from group 3 type
II and 1 from group 2 type I, were identified with reasonable sequence coverage from HPLC fraction 5, 7, 17, and 18. The identities
of the peptides attributed to selected gluten proteins were confirmed using MS/MS with BioMultiView to match the predicted
and measured partial amino acid sequences. Because of incomplete wheat DNA databases, many wheat gluten proteins could not
be identified. These results suggest that the combination of RP-HPLC with MS and MS/MS techniques is a promising approach
for the characterization of wheat gluten proteins. |
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