An on-line coupling of HPSEC-MALLS and a RP-HPLC procedure were used to characterize and to reveal the polydispersity of the glutenin polymers of doughs during mixing and resting. Experiments involved doughs prepared from several samples of a common French wheat cultivar (Soissons) differing in total amount of SDS-unextractable glutenin polymers. During dough mixing the amounts, the size distribution of protein and the glutenin subunit composition within the SDS-unextractable polymers changed. However, the major changes in SDS-unextractable glutenin content and size distribution occurred before the peak MT was reached, while detectable changes in subunit composition occurred also after the peak MT. Even if sonication, which was used to solubilize the total wheat glutenin, can narrow down the glutenin size distribution, HPSEC-MALLS revealed a close relationship between the SDS solubility of the glutenin polymers and their size distribution confirming a depolymerization and repolymerization hypothesis. During the depolymerization of the SDSunextractable polymers, glutenin subunits were released in nonrandom order, which was indicative of the polymers' having a hierarchical structure. Some HMW-GS (specially HMW-GS 1Dx5) were particularly resistant to the depolymerization mechanism. This suggested that the subunit plays a major role in forming the backbone of the SDS-unextractable polymers consistent with its potential to form branched structure. These studies suggest that the SDSunextractable polymers in flours have a well-ordered structure that can be modified by dough mixing and resting. Cereal ChemistryT. Aussenac et al. 3

Two varieties of wheat differing in high molecular weight glutenin subunit composition (Soissons, 5+10, Glu-D1a allele; Thésée, 2+12, Glu-D1a allele) were examined to follow the accumulation of polymeric proteins and the changes in molecular size distribution of these proteins during grain filling. The accumulation behaviour of polymeric proteins was determined by size-exclusion-HPLC, multistacking SDS-PAGE and the constituent polypeptides (high molecular weight and low molecular weight glutenin subunits) by reversed-phase-HPLC. For both cultivars, the accumulation of each class of protein was highly asynchronous, especially between the early deposition of SDS-soluble polymers and the late deposition of SDS-insoluble polymers, such that the average molecular size of polymeric protein increased in the period from 30 to 45 days after anthesis in natural conditions. By applying premature grain desiccations during the cell enlargement phase, it was demonstrated that the SDS-insoluble polymers formation was closely related with the process of water loss from the grain. Moreover, the rapid accumulation of SDS-insoluble polymers coincided with a rapid decrease in mass of both SDS-soluble polymers and monomers, suggesting an aggregative mechanism. Over the same period, the molecular size distribution of the polymers which can be used to differentiate the two genotypes studied, is highly correlated with the percentage of high molecular weight glutenin subunits in glutenins present in kernels when desiccation occurred. The formation of SDS-insoluble fraction is discussed in connection with the specific contribution of high molecular weight glutenin subunits to the formation of polymers (subunits linked by disulfide bonds).

No abstract

The accumulation of polymeric proteins and the changes in molecular size distribution of these proteins were followed during grain filling and/or premature desiccation. The accumulation behavior of polymeric proteins and their constituent polypeptides (high and low molecular weight glutenin subunits, HMW-GS and LMW-GS) was determined by reversed phase-high performance liquid chromatography using a NaI/propanol purification procedure. With this new extraction and separation procedure, we have demonstrated that there was a coordinated initiation of storage protein biosynthesis, even if the accumulation rate varied greatly between the two main classes of proteins (i.e. monomeric and polymeric fractions). Moreover, the glutenin subunit composition was largely modified during glutenin accumulation. Both the HMW-GS/LMW-GS and HMW-GS-x/HMW-GS-y ratios increased significantly during the whole cell enlargement phase (from 16 to 37 d after anthesis). By applying premature grain desiccation during this physiological phase, we demonstrated that the polymerization index (SDS-insoluble polymers/total polymers) of the glutenin polymers was closely related to the HMW-GS/LMW-GS ratio of these proteins. An increase in the relative proportion of HMW-GS in glutenins caused the proportion of SDS-insoluble polymers to rise during grain desiccation. From these studies, it appears that the modification of the desiccation rate (grain desiccation at a constant temperature with variable relative humidity levels) induced a parallel modification of the glutenin insolubilization rate but did not affect the polymerization index of the glutenins at maturity.

No abstract