Reoxidation Behavior of Wheat and Rye Glutelin Subunits

The ability of HMW and LMW subunits of wheat glutelin to form a polymeric gluten network by intermolecular disulfide bonds is responsible for the unique rheological properties and baking quality of wheat dough. Because the mechanism of gluten formation is not fully understood, the reoxidation behavior of HMW and LMW subunits of wheat glutelin and HMW subunits of rye glutelin was studied. The subunits were isolated from wheat flour cv. Rektor (REK) and from rye flour cv. Danko (DAN) with a selective extraction and precipitation method. For reoxidation, different oxidants (KBrO₃ and KIO₃), protein concentrations (0.5, 1.0, and 2.0%), solvent compositions, pH values (2.0 and 8.0), and reaction times (0–360 min) were compared. The characterization of reoxidized products was achieved by the determination of the thiol content with the Ellman's reagent, and of the M_r distribution by gel‐permeation chromatography. The results demonstrated that both HMW and LMW subunits could be slowly reoxidized with KBrO₃ to polymers with M_r up to several millions. Yield and M_r distribution of polymers were dependent both on the protein concentration and on the molar ratio of oxidants to thiol groups. The HMW subunits of wheat glutelin (HMW‐REK) yielded slightly higher quantities of polymeric proteins than did the HMW subunits of rye (HMW‐DAN). Reoxidation with KIO₃ proceeded much faster than with KBrO₃ and led to lower proportions of polymerized proteins for HMW‐REK and HMW‐DAN. Obviously, more intra‐ and fewer intermolecular disulfide bonds were formed by reoxidation with KIO₃ compared with KBrO₃. In contrast, LMW‐REK was reoxidized with KIO₃ to higher amounts of polymeric aggregates, which indicated that LMW subunits formed intermolecular disulfide bonds with both KIO₃ and KBrO₃. Independent of the protein type and the oxidant used for reoxidation, more inter‐ and fewer intramolecular disulfide bonds were formed when the protein concentration was increased. Single subunits 5, 7, and 10 were isolated from HMW‐REK by preparative acid‐PAGE and were reoxidized with KBrO₃ for 360 min. The M_r distribution indicated that x‐type subunit 5 had a greater tendency to form polymers than x‐type subunit 7. The y‐type subunit 10 was characterized by a lower proportion of polymers after reoxidation than x‐type subunits 5 and 7.     

Immunocytochemical Localization of Gluten Proteins Uncovers Structural Organization of Glutenin Macropolymer

The content and composition of the disulfide‐bonded glutenin macropolymer has been shown to influence dough properties, although its structural organization is poorly characterized. The structure of the glutenin macropolymer in dough was studied using an immunolocalization transmission electron microscopy (TEM) technique by localizing gliadins, low molecular weight glutenin subunits (LMW‐GS), and high molecular weight glutenin subunits (HMW‐GS) in sections of dough using antibody probes selective for each of the three classes of gluten polypeptides. Distinct differences in the distribution patterns of gliadins, LMW‐GS, and HMW‐GS were observed, which suggests that proteins have different roles in the structural organization of the gluten matrix. On the basis of the observed distribution of the proteins in dough, it is speculated that gliadins, which are randomly distributed as individual particles, fill space within the glutenin macropolymer; LMW‐GS, which are present as clusters, are speculated to form aggregated branch structures; and HMW‐GS, which are present as chains, are speculated to form a network from which the LMW‐GS branches are formed. Changes in the distribution of gliadins, LMW‐GS, and HMW‐GS in dough during mixing were also noted. Such an arrangement supports previous biochemical evidence which has established that gliadins, LMW‐GS, and HMW‐GS have specific roles in the structural organization of the glutenin macropolymer in doughs.     

Breadmaking Quality of Selected Durum Wheat Genotypes and Its Relationship with High Molecular Weight Glutenin Subunits Allelic Variation and Gluten Protein Polymeric Composition

Twenty‐seven durum wheat genotypes originating from different geographical areas, all expressing LMW‐2 at Glu‐B3, and five bread wheats were evaluated for flour mixing properties, dough physical characteristics, and baking performance. Gluten polymeric composition was studied using size‐exclusion HPLC of unreduced flour protein extracts. As a group, durum wheats had poorer baking quality than bread wheats in spite of higher protein and total polymer concentrations. Durum wheats exhibited weaker gluten characteristics, which could generally be attributed to a reduced proportion of SDS‐unextractable polymer, and produced less extensible doughs than did bread wheats. However, substantial variation in breadmaking quality attributes was observed among durum genotypes. Better baking performance was generally associated with greater dough extensibility and protein content, but not with gluten strength related parameters. Extensibility did not correlate with gluten strength or SEHPLC parameters. Genotypes expressing high molecular weight glutenin subunits (HMW‐GS) 6+8 exhibited better overall breadmaking quality compared with those expressing HMW‐GS 7+8 or 20. Whereas differences between genotypes expressing HMW‐GS 6+8 and those carrying HMW‐GS 7+8 could only be attributed to variations in extensibility, the generally inferior baking performance of the HMW‐GS 20 group relative to the HMW‐GS 6+8 group could be attributed to both weaker and less extensible gluten characteristics.     

In Vitro Polymerization of Wheat Glutenin Subunits with Inorganic Oxidizing Agents. I. Comparison of Single‐Step and Stepwise Oxidations of High Molecular Weight Glutenin Subunits

High molecular weight glutenin subunits (HMW‐GS) were isolated from wheat flour and polymerized in vitro at pH 3.0 with different oxidizing agents (KBrO₃, KIO₃, H₂O₂). An oxidation protocol with single addition of oxidant (single‐step oxidation) was compared with a set‐up in which the oxidant was added in multiple steps (stepwise oxidation). Changes in size distribution were evaluated with size‐exclusion HPLC, multilayer SDS‐PAGE, and flow‐field flow fractionation (flow‐FFF). Flow‐FFF is particularly suitable for measuring changes in glutenin size in the very high size ranges. In order of increasing sizes of the resulting polymers, the different oxidizing agents could be ranked as KBrO₃ < KIO₃ < H₂O₂. However, none of the oxidation conditions allowed for a complete polymerization of HMW‐GS. Interestingly, it was found that high concentrations of KIO₃ negatively affect the degree of polymerization. A similar observation was not made with KBrO₃ or H₂O₂. SDS‐PAGE showed that y‐type HMW‐GS particularly failed to incorporate in glutenin polymers. Simultaneously, these HMW‐GS displayed higher mobilities on SDS‐PAGE that can be ascribed to the formation of intrachain SS bonds. Possible explanations for the incomplete polymerization of HMW‐GS are given.     

Studies on Effects of Microbial Transglutaminase on Gluten Proteins of Wheat. I. Biochemical Analysis

The enzyme transglutaminase (TG) is known to have beneficial effects on breadmaking. However, only limited information is available on the structural changes of gluten proteins caused by TG treatment. The effect of TG has, therefore, been systematically studied by means of model peptides, suspensions of wheat flours and doughs. The treatment of synthetic peptides mimicking amino acid sequences of HMW subunits of glutenin with TG results in isopeptide bonds between glutamine and lysine residues. To study the effect on gluten proteins, different amounts of TG (0 to 900 mg enzyme protein per kg) were dissolved in a buffer and added to wheat flour. The flour suspensions were incubated and centrifuged and the residues were successively extracted with water, a salt solution, 60% aqueous ethanol (gliadin fraction) and SDS solution including a reducing agent (glutenin fraction). The characterization of the fractions by amino acid analysis, SDS‐PAGE, gel permeation HPLC and reversed‐phase HPLC has indicated that the quantity of extractable gliadins decreases by increasing TG amounts. Among gliadins, the ω5‐type was affected to the greatest extent by the reduction of extractability, followed by the ω1,2‐, α‐ and γ‐types. The oligomeric portion of the gliadin fractions (HMW gliadin) was strongly reduced when flour was treated with 450 and 900 mg TG per kg of flour, respectively. In the first instance, the quantity of the glutenin fractions increased by the treatment of flour with 90 and 450 mg TG per kg of flour, and significantly decreased by the treatment of flour with 900 mg TG per kg of flour. Parallel to an increase in TG concentration, the amounts of glutenin‐bound ω‐gliadins and HMW subunits were strongly reduced, whereas the LMW subunits reached a maximal amount after treatment with 450 mg TG per kg of flour. The insoluble residue was almost free of protein when flour was treated with lower amounts of TG. Higher amounts led to a great increase of protein in the residues. The effects of TG on doughs were similar to those of flour suspensions, but less strongly pronounced probably due to the lower water content of the dough system. Sequence analysis of peptides from a thermolytic digest of the insoluble residue revealed that HMW subunits of glutenin and α‐gliadins were predominantly involved in cross‐links formed by TG treatment.     

Biochemical Studies of Proteins in Nondeveloped,Partially Developed,and Developed Doughs

Nondeveloped, partially developed with shear and extensional deformations, and developed doughs represent different stages of dough development. To understand the relationship between gluten proteins and dough rheology, this study used disulfide‐sulfhydryl analyses, gel filtration chromatography, SDS‐PAGE, acid polyacrylamide gel electrophoresis (A‐PAGE), and densitometry to examine proteins in the four types of doughs mentioned. Free sulfhydryl content was the lowest in native flour and nondeveloped dough, and the highest in partially developed doughs, while a reverse trend was observed for disulfide content. For each flour sample, the protein elution profile from gel filtration chromatography shifted with the level of dough development. With respect to the smallest sized molecules, native flour had the most, followed by nondeveloped, partially developed, and then developed doughs. SDS‐PAGE and A‐PAGE exhibited similar protein patterns among the same chromatographed protein fractions of each native flour and its different doughs. Densitometric data showed that the amount of high molecular weight (HMW) glutenins increased and the amounts of low molecular weight (LMW) glutenins, gliadins, and albumins/globulins decreased with progressive stages of dough development. In conjunction with previously published results, indications are that the increase in the size and the amount of HMW glutenins is related to the strength of dough and the amount of protein matrix present in the dough.     

Effect of Multiple Substitution of Glutenin and Gliadin Proteins on Flour Quality of Canada Prairie Spring Wheat

The effect of genetic substitution of two to four glutenin and gliadin subunits from a Canada Prairie Spring (CPS) cv. Biggar BSR into Alpha 16, another CPS wheat line, was studied for rheological and baking quality. Results from double substitution showed that the presence of a gliadin component from Biggar BSR (BGGL) and low molecular weight glutenin subunit 45 (LMW 45) contributed to improved dough strength characteristics. Presence of BGGL in combination with high molecular weight glutenin subunit 1 (HMW 1) or 17+18 (HMW 17+18) also showed improved dough strength over control Alpha lines. When three or four protein subunits were substituted, even though improved quality performance was observed, it was associated with the negative effect of lowered flour water absorptions in spite of similar protein contents. The study confirms that LMW glutenins, as well as gliadins, play an important role along with HMW glutenins in wheat flour quality. CPS wheat lines with improved dough strength properties can be selected from the double substitution lines with the combination of BGGL/LMW 45 and BGGL/HMW 1.     

Glucose Oxidase in Breadmaking Systems

The mechanism of glucose oxidase action in breadmaking was investigated by studying the baking performance of glucose oxidase, the active ingredient that it produced, and its effect on the rheological properties of dough. Glucose oxidase improved the loaf volume of bread made by 45-, 70-, and 90-min fermentation processes. Although the increase in loaf volume was significant, it was less than that obtained with an optimum level of KBrO₃. With the 90-min fermentation process, the crumb grain of bread was similar for loaves oxidized with optimum levels of glucose oxidase or KBrO₃. The rheological properties of doughs containing glucose oxidase and doughs containing no oxidant were compared. Doughs made with glucose oxidase had higher G′ and G″ and lower tan δ values than doughs made without an oxidant. Hydrogen peroxide was responsible for a drying effect in doughs. This drying effect of glucose oxidase was reduced significantly by incorporation of free radical scavengers into the dough.     

Role of Gluten and Its Components in Determining Durum Semolina Dough Viscoelastic Properties

Gluten was isolated from three durum wheat cultivars with a range in strength. Gluten was further fractionated to yield gliadin, glutenin and high molecular weight (HMW) and low molecular weight (LMW) glutenin subunits (GS). The gluten and various fractions were used to enrich a base semolina. Enriched dough samples were prepared at a fixed protein content using a 2‐g micromixograph. Mixing strength increased with addition of gluten. Dynamic and creep compliance responses of doughs enriched with added gluten ranked in order according to the strength of the gluten source. Gliadin addition to dough resulted in weaker mixing curves. Gliadin was unable to form a network structure, having essentially no effect on dough compliance, but it did demonstrate its contribution to the viscous nature of dough (increased tan δ). Source of the gliadin made no difference in response of moduli or compliance. Addition of glutenin to the base semolina increased the overall dough strength properties. Glutenin source did influence both dynamic and compliance results, indicating there were qualitative differences in glutenin among the three cultivars. Enrichment with both HMW‐GS and LMW‐GS increased overall dough strength. Source of HMW‐GS did not affect compliance results; source of LMW‐GS, however, did have an effect. The LMW‐2 proteins strengthened dough to a greater extent than did LMW‐1. Mechanisms responsible for dough viscoelastic properties are described in terms of reversible physical cross‐links.     

Association of Glutenin Subunit Composition and Dough Rheological Characteristics with Cookie Baking Properties of Soft Wheat Cultivars

The effect of flour type and dough rheology on cookie development during baking was investigated using seven different soft winter wheat cultivars. Electrophoresis was used to determine the hydrolyzing effects of a commercial protease enzyme on gluten protein and to evaluate the relationships between protein composition and baking characteristics. The SDS‐PAGE technique differentiated flour cultivars based on the glutenin subunits pattern. Electrophoresis result showed that the protease degraded the glutenin subunits of flour gluten. Extensional viscosities of cookie dough at all three crosshead speeds were able to discriminate flour cultivar and correlated strongly and negatively to baking performance (P < 0.0001). The cookie doughs exhibited extensional strain hardening behavior and those values significantly correlated to baking characteristics. Of all rheological measurements calculated, dough consistency index exhibited the strongest correlation coefficient with baking parameters. The degradation effects of the protease enzyme resulted in more pronounced improvements on baking characteristics compared with dough rheological properties. Stepwise multiple regression showed that the dough consistency index, the presence or absence of the fourth (44 kDa) subunit in LMW‐GS and the fifth subunit (71 kDa) subunit in HMW‐GS were predominant parameters in predicting cookie baking properties.     

Depolymerization of the Glutenin Macropolymer During Mixing: II. Differences in Retention of Specific Glutenin Subunits

The depolymerization of individual high and low molecular weight (HMW and LMW, respectively) glutenin subunits (GS) from the glutenin macropolymer (GMP) in doughs during mixing was investigated by reversed-phase (RP) HPLC and SDS-PAGE. Cultivars with different dough strengths, as well as lines null for specific HMW-GS and biotypes differing at individual HMW-GS and LMW-GS encoding loci, were studied. During mixing, the proportion of total HMW-GS in GMP decreased, and the ratios of different subunits in the GMP in doughs changed. There was a loss of chromosome 1B- and 1D-encoded x-HMW-GS, while the relative proportions of y-HMW-GS (among HMW-GS) increased. Changes in 1B subunits occurred first, while most of the changes in 1D HMW-GS content occurred during dough breakdown. Changes were more pronounced for doughs of weak to average strengths than for stronger doughs. RP-HPLC analysis demonstrated a consistent increase in the retention times (surface hydrophobicity) of chromosome 1D-encoded HMW-GS but not of other HMW-GS or LMW-GS during mixing. SDS-PAGE and RP-HPLC demonstrated that specific B subunits, typically those with lower hydrophobicity, were selectively depolymerized from the GMP during dough breakdown, while the proportions of specific C subunits, typically those with greater hydrophobicity, increased. Similar trends were seen in analyses of several pairs of biotypes differing at single LMW-GS encoding loci, although there were slight differences in the depolymerization behavior of wheats with different allelic compositions. The results suggest that dough breakdown may be triggered by the loss of specific HMW-GS from the GMP, and a structural hierarchy may exist for different LMW-GS within glutenin in doughs.     

Simple Determination of Gluten Protein Types in Wheat Flour by Turbidimetry

A simple method based on turbidimetry has been developed for the quantitative determination of total gliadins, glutenin subunits, and high and low molecular weight (HMW and LMW) subunits of glutenin. The standard procedure includes the subsequent extraction of wheat flour (100 mg) with a salt solution, with 50% 2‐propanol (gliadins), and with 50% propanol under reducing conditions and increased temperature (glutenin subunits). Aliquots of the gliadin and the glutenin extracts are mixed with 2‐propanol to a final concentration of 83%, and the turbidity of the precipitates is measured photometrically at 450 nm and 20°C after 40 min. Another aliquot of the glutenin extract is mixed with acetone to a final concentration of 40% acetone, and precipitated HMW subunits are determined turbidimetrically after 30 min. The sample is then filtered, and an aliquot of the filtrate is mixed with 2‐propanol to a final concentration of 77% to determine the precipitated LMW subunits. Control analyses with reversed‐phase HPLC on C₈ silica gel indicate that the precipitation of the different protein types is quantitative and specific, and studies of 16 different wheat flours demonstrate the strong correlation between quantification by HPLC and turbidimetry. The turbidimetric measurements are reproducible, linear over a wide absorbance range (0.2–1.7), and sufficiently sensitive to analyze 40 μg of protein or 20 mg of flour. The absolute amounts of protein types in flour can be determined by means of calibration curves with protein standards (gliadins, HMW, and LMW subunits). Altogether, the developed method is simple, accurate, sensitive, and specific for the different protein types. The total procedure takes ≈6 hr for the analysis of six flour samples in parallel or ≈4 hr for three samples in overlapping extraction steps. The chemicals used are inexpensive, scarcely toxic, and easy to dispose.     

Quantitative Determination of Gluten Protein Types in Wheat Flour by Reversed-Phase High-Performance Liquid Chromatography

A combined extraction-HPLC procedure was developed on a microscale to determine the amounts of the different gluten protein types (ω5-, ω1,2-, α- and γ-gliadins; high molecular weight [HMW] and low molecular weight [LMW] glutenin subunits) in wheat flour. After preextraction of albumins and globulins from flour (100 mg) with a salt solution (2 × 1.0 mL), extraction of gliadins was achieved with 60% aqueous ethanol (3 × 0.5 mL). Subsequently, the glutenin subunits were extracted under nitrogen and at 60°C with 50% aqueous 1-propanol containing Tris-HCl (0.05 mol/L, pH 7.5), urea (2 mol/L) and dithioerythritol (1%). The separation and quantitative determination of gliadins and glutenin subunits was then performed by reversed-phase HPLC on C₈ silica gel at 50°C using a gradient of increasing acetonitrile concentration in the presence of 0.1% trifluoroacetic acid. The flow rate was 1.0 mL/min, and the detection wavelength was 210 nm. Temperature and flow rate were modified for the quantitation of single underivatized HMW subunits. To determine the absolute amounts of protein types, different protein standards (gliadin, LMW and HMW subunits, bovine serum albumin) with known protein contents were compared to HPLC absorbance areas. The calibration curves were almost identical and linear over a broad range (20–220 μg). This extraction-HPLC procedure allows an accurate, reproducible, sensitive, and relatively fast quantitative determination of all gluten protein types in wheat flour, and can be applied to quality evaluation of cereals as raw materials or in processed products.     

Expression,Purification, and Functional Analysis of Three Low‐Molecular‐Weight Glutenin Subunits from Wheat Cultivar Cheyenne

Glutenins, which form the network of gluten protein, are of great importance for the quality of flour products. Glutenins can be divided into HMW and LMW subunits according to molecular weight. Three genes for LMW glutenin subunits (LMW‐GS), named lmw‐cnd1, lmw‐cnd2, and lmw‐cnd3 with open reading frames of 1,053, 903, and 969 bp, respectively, were cloned from wheat cultivar Cheyenne. Heterologous expression vectors of the three LMW‐GS were constructed, and the recombinant proteins LMW‐CND1, LMW‐CND2, and LMW‐CND3 were overexpressed in Escherichia coli. After cell disruption with ultrasound, target proteins of high purity were obtained by using Ni²⁺ affinity chromatography. Farinograph and TAPlus measurements were used to investigate the effects of the three LMW‐GS on the characteristics of flour and dough. The results showed that the addition of each LMW‐GS can lead to an increase in the elasticity of the dough. Moreover, LMW‐CND2 and LMW‐CND3 promoted the strength of the dough. All three LMW‐GS caused a decrease of hardness and increase of springiness and cohesiveness of dough according to texture profiling results. Consequently, all three LMW‐GS have positive effects on the processing characteristics of dough and can improve bread quality to different extents.     

Characterization of Monomeric and Glutenin Polymeric Proteins of Hard Red Spring Wheats During Grain Development by Multistacking SDS-PAGE and Capillary Zone Electrophoresis

Three cultivars of hard red spring (HRS) wheats with identical high molecular weight (HMW) glutenin subunit composition (5+10 type, Glu-D1d) but different dough properties and breadmaking quality were used in this study. The synthesis and accumulation characteristics of different protein fractions during grain development were examined. Samples were collected at three-day intervals from anthesis to maturity between day 10 to day 37. The nonreduced SDS-extractable glutenin aggregates of developing grains were characterized by a multistacking SDS-PAGE procedure to obtain information on the size distribution and polymerization of glutenin aggregates. The HMW to low molecular weight (LMW) glutenin subunit ratio was determined for its relationship to polymerization of the various glutenin aggregates of different molecular sizes. Glutenin proteins were quantified using an imaging densitometer. In addition, albumins and globulins, α- and β-gliadins, γ-gliadins, and ω-gliadins were separated by capillary zone electrophoresis. The results indicated that albumins-globulins, gliadins, and glutenins in developing grains were present at 10 days after anthesis or earlier. Albumin-globulins decreased in proportion, while gliadins increased in proportion during grain development. Polymerization of glutenin aggregates occurred 10 days after anthesis or earlier and increased significantly throughout the grain-filling period until maturity. Larger aggregates of glutenin increased in proportion, while smaller ones decreased in proportion during grain development. Ratio of polymers to monomers increased significantly from day 10 to day 22 of grain development and then remained constant until grain maturity. Glutenin polymers arrived at their maximum in proportion to total SDS-extractable proteins or monomers at day 22 after anthesis while the molecular size of these polymers continued to increase, as indicated by a rapid increase in proportion of HMW to LMW glutenin subunits. Significant differences were found in accumulation rates of glutenin polymers among the three cultivars. Cultivars Kulm and Grandin, with better breadmaking quality, appeared to have greater rates of accumulation and HMW subunit synthesis or formation of larger polymers than did Sharp, a cultivar with poorer quality. Significant differences were found among the three cultivars in the proportion of albumins-globulins and gliadins during grain development. However, no significant differences were found among the cultivars in the proportion of albumins-globulins, α-, β-, γ-, and ω-gliadins at grain maturity. Varietal differences in breadmaking quality were due mainly to the differences in glutenin polymers such as ratio of polymeric to monomeric proteins, molecular size distribution, and ratio of HMW to LMW glutenin subunits among wheat cultivars of 2*, 7+9, and 5+10 subunit types. The better breadmaking cultivars might be characterized with higher proportions of glutenins and greater proportion of HMW subunits in total SDS-extractable proteins than the poorer quality cultivar. However, more genotypes need to be examined.     

Influence of High Molecular Weight (HMW) and Low Molecular Weight (LMW) Glutenin Subunits Controlled by Glu‐1 and Glu‐3 Loci on Durum Wheat Quality

The progenies of four intervarietal durum wheat crosses were used to determine the effects of glutenin variants coded at Glu‐1 and Glu‐3 loci on durum wheat quality properties. The F₂ lines were analyzed for high molecular weight (HMW) and low molecular weight (LMW) glutenin composition by electrophoresis. Whole grain derived F₃ and F₄ samples were analyzed for vitreousness, protein, and dry gluten contents, gluten index, SDS sedimentation volume, mixograph, and alveograph properties. Allelic variation at the Glu‐B1 and Glu‐B3 loci affected gluten quality significantly. Comparisons among the Glu‐B3 and Glu‐B1 loci indicated that the LMW glutenin subunits controlled by Glu‐B3 c and j made the largest positive contribution, followed by the alleles a, k, and b. HMW glutenin subunits 14+15 gave larger SDS values and higher mixing development times than subunits 7+8 and 20. The positive effects of the glutenin subunits LMW c and HMW 14+15 were additive. Flour protein content, vitreousness, and mixograph peak height values were positively correlated with each other as well as with Dglut values, whereas the SDS sedimentation highly correlated with mixing development time, alveograph strength, and extensibility but was not correlated with the other parameters. The results of quality analysis, together with the results of the genetic analysis, led to the conclusion that SDS sedimentation, mixograph mixing development time, and peak breakdown are the tests more influenced by allelic variation of prolamin. The uses of the results in durum wheat quality breeding programs are discussed.     

Preparative Method for In Vitro Production of Functional Polymers from Glutenin Subunits of Wheat

An in vitro method for preparative‐scale production of artificial glutenin polymers utilizes a controlled environment for the oxidation of glutenin subunits (GS) isolated from wheat flour to achieve high polymerization efficiency. The functionality of in vitro polymers was tested in a 2‐g model dough system and was related to the treatment of the proteins before, during, and after in vitro polymerization. When added as the only polymeric component in a reconstituted model dough (built up from gliadin, water solubles, and starch fractions), in vitro polymers could mimic the behavior of native glutenin, demonstrating properties of dough development and breakdown. Manipulating the high molecular weight (HMW)‐GS to a low molecular weight (LMW)‐GS ratio altered the molecular weight distribution of in vitro polymers. In functional studies using the 2‐g mixograph, simple doughs built up from homopolymers of HMW‐GS were stronger than those using homopolymers of LMW‐GS. These differences may be accounted for, at least in part, by different polymer size distributions. The ability to control the size and composition of glutenin polymers shows the potential of this approach for investigating the effects of glutenin polymer size on dough function and flour end‐use quality.     

Composition of HMW and LMW Glutenin Subunits and Their Effects on Dough Properties,Pan Bread,and Noodle Quality of Chinese Bread Wheats

Knowledge of composition of high molecular weight glutenin subunits (HMW‐GS) and low molecular weight glutenin subunits (LMW‐GS) and their associations with pan bread and noodle quality will contribute to genetically improving processing quality of Chinese bread wheats. Two trials including a total of 158 winter and facultative cultivars and advanced lines were conducted to detect the allelic variation at Glu‐1 and Glu‐3 loci by SDS‐PAGE electrophoresis and to understand their effects on dough properties, pan bread, and dry white Chinese noodle (DWCN) quality. Results indicate that subunits/alleles 1 and null at Glu‐A1, 7+8 and 7+9 at Glu‐B1, 2+12 and 5+10 at Glu‐D1, alleles a and d at Glu‐A3, and alleles j and d at Glu‐B3 predominate in Chinese germplasm, and that 34.9% of the tested genotypes carry the 1B/1R translocation (allelic variation at Glu‐D3 was not determined because no significant effects were reported previously). Both variations at HMW‐GS and LMW‐GS/alleles and loci interactions contribute to dough properties and processing quality. For dough strength related traits such as farinograph stability and extensigraph maximum resistance and loaf volume, subunits/alleles 1, 7+8, 5+10, and Glu‐A3d are significantly better than those of their counterpart allelic variation, however, no significant difference was observed for the effects of d, b, and f at Glu‐B3 on these traits. For extensigraph extensibility, only subunits 1 and 7+8 are significantly better than their counterpart alleles, and alleles d and b at Glu‐B3 are slightly better than others. For DWCN quality, no significant difference is observed for HMW‐GS at Glu‐1, and Glu‐A3d and Glu‐B3d are slightly better than other alleles. Glu‐B3j, associated the 1B/1R translocation, has a strong negative effect on all quality traits except protein content. It is recommended that selection for subunits/alleles 1, 7+8, 5+10, and Glu‐A3d could contribute to improving gluten quality and pan bread quality. Reducing the frequency of the 1B/1R translocation will be crucial to wheat quality improvement in China.     

Comparative study of the effect of incorporated individual wheat storage proteins on mixing properties of rice and wheat doughs

The aim of this work was to compare the effects of incorporated wheat storage proteins on the functional properties of rice and wheat flours. The advantage of rice as a base flour compared to wheat is that it does not contain any wheat flour components and, therefore, has no interactive effect between wheat glutenin proteins. The incorporation of individual HMW glutenin subunit proteins (Bx6, Bx7, and By8) in different ratios had significant positive effects on the mixing requirements of both rice and wheat doughs. Reconstitution experiments using two x+y type HMW-GS pairs together with a bacterially expressed LMW-GS have been also carried out in this study. The largest effects of polymer formation and mixing properties of rice flour dough were observed when Bx and By subunits were used in a 1:1 ratio and HMW and LMW glutenin subunits in a 1:3 ratio. However, using the same subunit ratios in wheat as the base flour, these synergistic effects were not observed.     

Relationship Between the Qualitative and Quantitative Compositions of Gluten Protein Types and Technological Properties of Synthetic Hexaploid Wheat Derived from Triticum durum and Aegilops tauschii

The contribution of the diploid wheat species Aegilops tauschii (Coss.) Schmall to the technological properties of bread wheat (Triticum aestivum L.) was previously studied by the investigation of synthetic hexaploids derived from tetraploid durum wheat (T. turgidum L.) and three diploid Ae. tauschii lines. The results indicated that bread volume, gluten index, SDS‐sedimentation volume, and maximum resistance of gluten were significantly influenced by the Ae. tauschii lines. To determine the relationship between technological properties and qualitative and quantitative compositions of gluten proteins, the flours of parental and synthetic lines were extracted using a modified Osborne fractionation. Gliadin and glutenin fractions were then characterized by reversed‐phase (RP) HPLC on C₈ silica gel. The HPLC patterns revealed typical differences between synthetic and parental lines. The gliadin patterns of three synthetic lines and the glutenin patterns of two synthetic lines were more similar to that of the diploid Ae. tauschii parents involved in the hybrids. In the glutenin pattern of one synthetic line, characteristics from both Ae. tauschii and the durum wheat parents were observed. The amount of total gliadin and gliadin types of the synthetic lines was mostly intermediate between those of the durum and Ae. tauschii parents. The amounts of total glutenin and glutenin types (HMW and LMW subunits) of the synthetic lines were generally higher than those of the parental lines, and the ratio of gliadins to glutenins was significantly decreased. High positive correlations were found between the amount of total glutenins, HMW, and LMW subunits and bread volume, maximum resistance and extension area of gluten, and SDS‐sedimentation volume. The ratio of gliadins to glutenin subunits had a strong negative influence on these properties. The protein content of the flours and the amount of total gluten proteins were not correlated with any of the technological properties. Results on the relationship between biochemical characteristics and the breadmaking properties indicated that wheat prebreeding would benefit from studies on protein types and quantification in the choice of parents. In addition, the potential of the diploid Ae. tauschii for improvement of breadmaking quality should be further exploited.