Location: Crop Improvement and Genetics Research
Project Number: 5325-43000-027-00-D
Project Type: In-House Appropriated
Start Date: Oct 1, 2010
End Date: Sep 24, 2013
The first objective of the research is to determine the basis for changes in flour quality that result from high temperatures during wheat grain development. The research will investigate the roles of gluten composition and polymer structure in effects of temperature during grain development on flour quality. The research also will determine the roles of specific non-gluten proteins in effects of temperature during grain development on flour quality. This objective addresses a major concern of millers and bakers and explores two hypotheses: 1) changes in glutenin polymer amount, size, structure and composition as a result of high temperatures during grain development are responsible for decreases in flour quality and 2) non-gluten proteins that increase in the grain under high temperature conditions are involved in quality or allergenicity. The second objective of the research is to identify and characterize wheat proteins responsible for human intolerances and allergies that affect nearly 2% of the U.S. population and to develop methods to detect allergenic proteins in downstream products. This objective will determine whether mass spectrometry (MS) can be used to identify potential wheat allergens in flour and detect these proteins in food ingredients and products.
To address the first objective, MS methods coupled with improved methods for protease digestion will be developed so that closely related gluten proteins can be distinguished. Size-exclusion (SE) chromatography and high pressure liquid chromatography (HPLC) will be used to separate glutenin polymers into size classes for determination of subunit composition and key linkages between high molecular weight glutenin subunits (HMW-GS) and low molecular weight glutenin subunits (LMW-GS). Gluten proteins that act as chain terminators in polymer structure will be identified and their roles in polymer structure and size will be evaluated. Polymer composition and size will be measured during grain development under different temperature regimens. The effect of different temperature regimens on accumulation profiles of a specific set of non-gluten proteins and their transcripts during grain development also will be characterized using 2-dimensional polyacrylamide gel electrophoresis (2-DE) and quantitative reverse transcriptase polymerase chain reaction (QRT-PCR). Since many of these proteins may be involved in stabilizing gas bubbles in dough, experiments will be performed to test whether the levels of these proteins increase in dough liquor prepared from flour from grain grown under high temperatures. Tissue localization studies will be performed and the roles that specific proteins play in flour quality and allergenicity will be investigated using transgenic plants in which the corresponding genes are suppressed. To address the second objective, the allergenic potential of non-gluten proteins that increase under high temperature conditions will be tested using sera from patients with defined wheat allergies. MS will be used to determine mass profiles of protein fractions from wheat flour. These profiles will be examined for signatures of specific allergenic proteins. Methods will be extended to samples from baked products such as bread. Replacing 5325-43000-026-OOD (June/2010).