Characterization and Quantification of Native Glutenin Aggregates by Multistacking Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS‐PAGE) Procedures

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Abstract
Native glutenin aggregates of two different quality flours containing the same high molecular weight (HMW) glutenin subunit compositions were investigated by multistacking sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) procedures. Five stacking gels (4, 6, 8, 10, and 12%) with a separating (resolving) gel of 14% were used to separate nonreduced glutenin aggregates solubilized from flour by SDS sodium phosphate buffer. There were large differences in protein solubility of the two flours. It took 8 hr to extract 91% of total proteins from the strong flour (variety Len) while it took only 2 hr for the weak flour sample 205. Total glutenin proteins and the proportions of glutenins at the different origins (including the origin of the 14% separating gel) were quantified by high‐resolution densitometry procedures. As the duration of extraction increased, both total glutenins and glutenins at the 4% origins increased. The good quality flour Len had a higher total glutenin protein (3% more) and higher proportion of glutenins with the largest molecular sizes (also 3% more) at the 4% origins than the poor quality flour sample 205. After glutenin aggregates from each origin were reduced and analyzed by SDS‐PAGE, the largest glutenins at the 4% origin contained twice the amount of total HMW glutenin subunits when compared to the smaller aggregates at the 12% origin. Among the total HMW glutenin subunits, the proportion of subunit 5 (which published literature reports to be the largest molecular weight based on calculations of DNA‐derived amino acid sequence analysis) was twice that at the 12% origin. A randomized structure of native glutenins is proposed based on the results of our investigations.