High Molecular Glutenin Subunits Research Articles

Molecular Characterization and Marker Development of the HMW-GS Gene from Thinopyrum elongatum for Improving Wheat Quality

The quality of wheat primarily depends on its storage protein quality, especially in regards to gluten content and high-molecular-weight glutenin subunits (HMW-GS). The number of HMW-GS alleles is limited in bread wheat (Triticum aestivum L.), whereas it is abundant in wheat relatives. Therefore, HMW-GS alleles from wheat relatives could provide a potential for improving quality in wheat breeding. Thinopyrum elongatum (EE) is one of the relatives of wheat. The E genome is closely related to the ABD genome in wheat; therefore, Th. elongatum is often used as an excellent exogenous gene donor for wheat genetic improvement. In this study, the high-molecular glutenin subunit gene was cloned and sequenced from Th. elongatum. A specific molecular marker for identifying the Glu-1Ey subunit gene was developed and applied to detected wheat-Th. elongatum alien introgression lines. Quality analysis indicated that the substitution and addition lines containing Th. elongatum alleles significantly (p < 0.05) increased grain protein content by 3.76% to 5.11%, wet-gluten content by 6.55% to 8.73%, flour 8-MW by 0.25% to 6.35%, and bread volume value by 33.77 mL to 246.50 mL, in comparing it with Chinese Spring. The GMP content and lactic acid SRC showed significant positive correlations with flour processing quality and might be used as indicators for wheat quality. The results were expected to provide a novel route for improving processing quality in wheat quality breeding.

Conformational rearrangement and polymerization behavior of frozen-stored gluten during thermal treatment

To elucidate the underlying mechanism of thermal-induced polymerization process of frozen-stored gluten, the conformational variations and polymerization behavior of fresh and frozen-stored gluten fractions during thermal treatment were comparatively tracked. The combined results of Raman spectra and fluorescence quenching by acrylamide suggested that frozen-stored gluten exhibited lower sensitivity to unfolding response upon heating, and more exposed tryptophan rather than tyrosine was detected for frozen-stored gluten when exceeding 70 °C, leading to the higher apparent binding constant for acrylamide and tryptophan than that of fresh gluten consequently. After the polymerization of glutenin and gliadin at 95 °C, the unstable trans-gauche-trans (t-g-t) conformation of disulfide bonds for frozen-stored gluten occupied more than 80%, while the stable gauche-gauche-gauche (g-g-g) diminished. The trans-gauche-gauche (t-g-g), t-g-t and g-g-g contents for the fresh gluten were similar, accounting for nearly 30%, respectively. Confocal laser scanning microscopy revealed the distorted glutenin network and heterogeneously distributed gliadin aggregates in frozen-stored gluten during thermal treatment. The polymerization ability of high molecular glutenin subunit Ax2, 1Dx5, Bx7.1 as well as the α- and γ-gliadin fractions in frozen-stored gluten were weakened compared with the fresh one.

Molecular characterization of high-molecular weight glutenin subunits 1Bx6.1 and 1By22.1 from Triticum spelta К1731 accession

Aim. Some spelt varieties, along with alleles of gliadins and high-molecular glutenin subunits (HMW-GS), identical to common wheat, contain specific alleles, that are source of Triticum aestivum gene pool enrichment. The aim of this work is the identification, molecular analysis of HMW – GS from T. spelta K1731 and evaluation of their effect on the elastic properties of gluten. Methods. Identification of HMW-GS was carried out by SDS-electrophoresis and PCR analysis. Nucleotide gene sequences were determined by Sanger sequencing. The secondary structure of proteins was predicted on the on-line CFSSP server. Results. Subunits 6.1 + 22.1 of the Glu-B1 locus encoded by the Glu-B1be allele were detected in the T. spelta K1731. The nucleotide sequences of the 1Bx6.1, 1By22.1 genes from spelt were determined, the amino acid sequence and the protein secondary structure of 6.1 + 22.1 subunits were analyzed. Conclusions. Molecular analysis of HMW-GS 1Bх6.1 and 1By22.1 from T. spelta К1731 established a low contribution to the bread-making quality of these subunits.
 Keywords: Triticum spelta K1731, HMW-GS, SDS electrophoresis, sequencing, secondary protein structure, gluten quality.

Molecular characterization and phylogenetic analysis of unusual x-type HMW glutenin subunits from 1sl genome of Aegilops longissima

Wheat related diploid species Ae. longissima (2n=2x=14, SlSl) has extensive storage protein variations that may provide useful gene resources for wheat quality improvement. In this work, five novel 1Sl-encoded x-type high molecular glutenin subunits (HMW-GS) were identified and designated as 1Slx-123, 1Slx-129, 1Slx1-136, 1Slx2-136 and 1Slx2.2, respectively. Their complete open reading frames (ORFs) were cloned and sequenced by AS-PCR, which contained 2874 bp (956 aa) for 1Slx-123, 2946 bp (979 aa) for 1Slx-129, 2901 bp (965 aa) for 1Slx1-136, 2982bp (991 aa) for 1Slx2-136 and 2928 bp (974 aa) for 1Slx2.2. Molecular characterization demonstrated that five unusual subunits had greater repetitive domains resulted from a larger fragment insertion (74-113 aa). Particularly, 1Slx-129 had an extra cysteine residue at the position 109 due to a TAT ? TGT dot mutation, which may improve the formation of superior gluten macropolymer. Our results suggest that these unusual HMW-GS could be served as potential superior gene resources for improving wheat gluten quality. Phylogenetic analysis revealed that HMW-GS genes from Glu-1Sx genomes had close evolutionary relationships with those of Glu-1Dx genome while sequences from Ae. speltoides aligned with those of B genome.

In vitro Functional Testing of High Molecular Glutenin Subunits in Wheat

The functions of high molecular weight glutenin subunit(HMW-GS) can be analyzed by determining the parameters subject to wheat(Triticum aestivum L.) quality.However,to date,only several glutenin subunits have verified in their functions despite a lot of subunits designated.Therefore,a simple and rapid method is highly needed to detect the functions of wheat glu-tenin subunits.The 2 g and 10 g Mixograph have been applied in related studies in Australia and other countries.To determine the function of wheat single HMW-GS,6 subunits,1Ax1,1Dx5,1Dy10,1Bx7,1By8,and 1By9,were separated and purified by SDS-PAGE,and then mixed into flour of Jing 411 individually.Their effects on flour mixing properties were investigated by 10 g Mixograph testing.The tested mixing parameters were mixing time,stability time,peak resistance,peak width,right slop,and width at 8 min.Four kinds of solution were applied to obtain an appropriate control,and the solutions consisted of different com-binations of 0.2% acetic acid,50 μg mL-1 DTT,200 μg mL-1 KIO3,and 0.1% SDS.When signal subunit was added into dough,6 mixing parameters changed to a different degree.The effects of the 6 subunits on flour quality were ranked as 1Dy10 1Ax1 1Dx5 1By8 1Bx7 1By9 according to mixing time,and 1Ax1 and 1Dy10 1Dx5 1By9 1Bx7 1By8 according to sta-bility time.The peak resistance of 1By8 was the biggest among that of all subunits.The right slop varied little among that of the 6 subunits.The width at 8 min of 1Bx7 was the smallest among that of all subunits.The 1By8 was primarily considered as a good quality subunit in this study.In addition,it is suggest that in vitro small-scale testing used in this study is an effective method to study the function of single HMW subunit.

High-Molecular-Weight Glutenin Subunit Composition of Chinese Wheat Germplasm

Abstract The objective of the present study was to characterize the high molecular glutenin subunits (HMW-GS) composition and the presence of 1B/1R translocation in newly developed wheat ( Triticum aestivum L.) germplasm, which have one or more traits that are useful in wheat improvement. Sodium dodecyl sulphate polyacrylamide-gel electrophoresis (SDS-PAGE) and acid polyacrylamide-gel electrophoresis (A-PAGE) were used to detect HMW-GS composition and the presence of 1B/1R wheat-rye ( Secale cereale L.) chromosome translocation in the wheat germplasm. Bread-making quality scores of these lines were determined. A high level of variations in HMW-GS encoded by Glu-1 locus was observed. Sixteen major HMW-GS, with 30 combinations, were detected. The percentage of cultivars with more than two desirable subunits was 38.7%. Thirteen cultivars had bread-making quality scores of 10 in combination with one or two desirable agronomical traits, such as high-yield potential, dwarfing stem, resistance to diseases, and/or tolerance to abiotic stress. Sixty-eight (36.6%) cultivars possessed 1B/1R translocation. The newly developed germplasm with HMW-GS for good quality can be promising resources for improving bread-making quality of wheat.

Size Characterisation of Glutenin Polymers by HPSEC-MALLS

The use of an on-line coupling of a new high-performance size exclusion chromatography (HPSEC) phase characterised by a very high exclusion limit and a multiangle laser light scattering detection (MALLS) for the size characterisation of wheat glutenin polymer was examined. Flour glutenin polymers which were extracted and purified by differential solubility in aqueous 50 and 70% propan-1-ol were solubilised with or without sonication in combination with a surfactant, 2% SDS. The glutenin polymers of the two genotypes studied, Soissons (5+10, Glu-1D) and Thésée (2+12, Glu-1D), were characterised by large polydispersity and by significant differences of size distribution of total polymers. The molecular size distribution of the total polymers, which can be used to differentiate the two genotypes, was highly correlated with the percentage of high-molecular glutenin subunits (HMW-GS) in glutenins. Furthermore, the results obtained for both the SDS-soluble and SDS-insoluble glutenin polymers from the two varieties revealed significant differences in size distribution, and also in molecular conformation (dependence of size upon mass). The molecular dimensions of SDS-insoluble polymers increased more slowly with the molecular weight than SDS-soluble polymers, which suggested a more compact structure. To conclude, the method appears to be a viable procedure to obtain rapid size characterisation of unreduced glutenin.

Recent advances in structure and function of food proteins: QSAR approach.

QSAR (quantitative structure-activity relationship), widely used in chemistry with hydrophobic, electronic, and steric parameters as structural factors, was found to be appropriate for use with food proteins, despite the difficulty, due to the complexity in macromolecular structure, in defining the steric terms. Emulsifying ability was closely related to hydrophobicity, and incorporation of solubility to hydrophobicity as factors improved the R2 of regression analysis. Foaming activity required both hydrophobicity and other factors pertaining to the adsorption of proteins at the interface in order to obtain adequate foam lamella strength. Hydrophobicity as well as other factors relating to the intermolecular interactions, for example, Ca and SH are involved in thermally induced gelation. For breadmaking, although no extensive QSAR work had been carried out, the important function of high molecular glutenin subunits was confirmed, and, notably, the critical function of hydrophobicity in breadmaking also was demonstrated. PLS (partial least-squares regression) and neural networks classify more correctly than other multivariate techniques, thereby yielding higher r2 values in modeling and prediction. However, multiple regression analysis and PCR (principal component regression) also were found to be effective for modeling because the information useful in elucidating the mechanism of protein function could be readily obtained. A characteristic property of unsupervised learning techniques, especially PCS (principal component similarity analysis), in identifying influential factors in the function mechanisms was demonstrated.