The structure and properties of gluten: an elastic protein from wheat grain

The wheat gluten proteins correspond to the major storage proteins that are deposited in the starchy endosperm cells of the developing grain. These form a continuous proteinaceous matrix in the cells of the mature dry grain and are brought together to form a continuous viscoelastic network when flour is mixed with water to form dough. These viscoelastic properties underpin the utilization of wheat to give bread and other processed foods. One group of gluten proteins, the HMM subunits of glutenin, is particularly important in conferring high levels of elasticity (i.e. dough strength). These proteins are present in HMM polymers that are stabilized by disulphide bonds and are considered to form the 'elastic backbone' of gluten. However, the glutamine-rich repetitive sequences that comprise the central parts of the HMM subunits also form extensive arrays of interchain hydrogen bonds that may contribute to the elastic properties via a 'loop and train' mechanism. Genetic engineering can be used to manipulate the amount and composition of the HMM subunits, leading to either increased dough strength or to more drastic changes in gluten structure and properties.     

Effects of cell wall components on the functionality of wheat gluten

Normal white wheat flours and especially whole meal flour contain solids from the inner endosperm cell walls, from germ, aleurone layer and the outer layers of cereal grains. These solids can prevent either gluten formation or gas cell structure. The addition of small amounts of pericarp layers (1–2%) to wheat flour had a marked detrimental effect on loaf volume. Microstructural studies indicated that in particular the epicarp hairs appeared to disturb the gas cell structure. The detrimental effects of insoluble cell walls can be prevented by using endoxylanases. It has been shown that some oxidative enzymes, naturally present in flour or added to the dough, will oxidise water-extractable arabinoxylans via ferulic acid bridges, and the resulting arabinoxylan gel will hinder gluten formation. The negative effects of water-unextractable arabinoxylans on gluten yield and rheological properties can be compensated by the addition of ferulic acid. Free ferulic acid can probably prevent arabinoxylan cross-linking via ferulic acid.     

Down-regulating γ-gliadins in bread wheat leads to non-specific increases in other gluten proteins and has no major effect on dough gluten strength

Background

Gliadins are a major component of gluten proteins but their role in the mixing of dough is not well understood because their contribution to wheat flour functional properties are not as clear as for the glutenin fraction.

Methodology/Principal Findings

Transgenic lines of bread wheat with γ-gliadins suppressed by RNAi are reported. The effects on the gluten protein composition and on technological properties of flour were analyzed by RP-HPLC, by sodium dodecyl sulfate sedimentation (SDSS) test and by Mixograph analysis. The silencing of γ-gliadins by RNAi in wheat lines results in an increase in content of all other gluten proteins. Despite the gluten proteins compensation, in silico analysis of amino acid content showed no difference in the γ-gliadins silenced lines. The SDSS test and Mixograph parameters were slightly affected by the suppression of γ-gliadins.

Conclusions/Significance

Therefore, it is concluded that γ-gliadins do not have an essential functional contribution to the bread-making quality of wheat dough, and their role can be replaced by other gluten proteins.     

Rheological and breadmaking properties of wheat samples infected with<Emphasis Type="Italic">Fusarium spp.</Emphasis>

Rheological and breadmaking properties of untreated and suboptimally stored wheat samples (grain moisture: 20%, temperature: 20°C) and also of wheat which was inoculated withFusarium spp. were investigated. The deoxynivalenol (DON) content of the stored and inoculated wheat samples ranged between 820–12,000 μg/kg. Gluten proteins were isolated with different extraction solutions and the fractions obtained were analysed by means of RP-HPLC. Microextension tests and micro-baking tests were used for the determination of dough properties (maximum resistance (MR) and extensibility (EX)) and bread volume, respectively. In spite of the extremely high DON concentrations of some wheat samples contaminated withFusarium spp. they showed only a slight decrease of the amount of gluten proteins. Extension tests of dough led to a slight decrease of MR, bread volumes stayed almost the same compared with the non-contaminated grain. The contamination of wheat withAspergillus andPenicillium led to a high decrease of gluten proteins, which resulted in an extremely decreased MR of the dough and a very low bread volume.     

Effect of arabinogalactan isolated from Siberian larch on the baking value of soft wheat flour and bread quality

Using additives of arabinogalactan (AG) isolated from Siberian larch, we examined the soft wheat flour quality and quantity of gluten, physical properties of the dough, and quality of finished bread depending on the quantity of the added polysaccharide. In the case of the addition of 1–3% of AG to flour, its content decreases in the final product. An excess amount of AG inhibits yeast growth, which leads to a decrease in bread quality. The optimum addition of AG to flour is 1%, at which the technological properties of flour and dough do not change significantly, but the quality of bread becomes remarkably better; furthermore, arabinogalactan is fully consumed in the course of bread preparation. The use of AG is recommended in the optimum dose for increasing the quality of baked goods.     

Viscoelastic properties of wheat gluten in a molecular dynamics study

Wheat (Triticum spp.) gluten consists mainly of intrinsincally disordered storage proteins (glutenins and gliadins) that can form megadalton-sized networks. These networks are responsible for the unique viscoelastic properties of wheat dough and affect the quality of bread. These properties have not yet been studied by molecular level simulations. Here, we use a newly developed α-C-based coarse-grained model to study ∼ 4000-residue systems. The corresponding time-dependent properties are studied through shear and axial deformations. We measure the response force to the deformation, the number of entanglements and cavities, the mobility of residues, the number of the inter-chain bonds, etc. Glutenins are shown to influence the mechanics of gluten much more than gliadins. Our simulations are consistent with the existing ideas about gluten elasticity and emphasize the role of entanglements and hydrogen bonding. We also demonstrate that the storage proteins in maize and rice lead to weaker elasticity which points to the unique properties of wheat gluten.     

花后灌水次数对强筋小麦籽粒产量和品质的影响

在防雨池栽培条件下,以2个优质强筋小麦品种（济麦20和藁城8901）为试验材料,研究了花后不同水分供应状况对籽粒产量、籽粒品质（粉质仪参数和面包体积）及其蛋白质组分的影响.结果表明：2个品种的籽粒产量、面粉的面团形成时间、面团稳定时间和制成面包体积均随花后灌水次数的增加呈先升高后降低的趋势;其中,花后灌1水（花后7 d）时藁城8901的籽粒产量最高,花后灌2水（花后7 d+花后14 d）时济麦20的籽粒产量最高;2个品种面粉的面团形成时间、面团稳定时间和制成面包体积均以花后灌1水时最优.单体蛋白含量、不溶性谷蛋白含量、谷蛋白总含量、蛋白质含量以及湿面筋含量也呈现类似的变化趋势.逐步回归分析表明,花后不同水分供应状况下,不溶性谷蛋白含量是影响面团稳定时间的关键因素,谷蛋白总含量与面包体积的变化密切相关.因此,为了保持优质强筋小麦品质的稳定性,水分管理应以改善籽粒蛋白质特别是谷蛋白组分的构成为目标.     

花后灌溉对小麦籽粒贮藏蛋白聚合程度和面团流变学特性的影响

为了明确灌溉麦黄水引起的小麦(Triticum aestivum)面团流变学特性劣变与籽粒贮藏蛋白聚合程度变化之间的关系, 在防雨池栽条件下, 以‘济麦20’为供试品种, 设置花后不灌水(W0), 花后灌1水(花后14 d, W1)和花后灌2水(花后14 d和花后28 d, W2) 3个处理, 分析了花后灌水对籽粒产量、贮藏蛋白聚合程度相关参数及其面团流变学特性的影响。研究结果表明, 籽粒产量、面团形成时间和稳定时间均以花后灌1水时达到最优, 再增加一次灌水(麦黄水), 导致面团形成时间和稳定时间显著缩短, 筋力变弱。同时观察到谷蛋白聚合指数和谷蛋白大聚合体平均粒径也因灌麦黄水而显著降低。回归分析表明, 灌麦黄水条件下谷蛋白大聚合体粒径变小是导致面团流变学特性变差和筋力变弱的主要原因。     

Xyloglucan oligosaccharides promote growth and activate cellulase: evidence for a role of cellulase in cell expansion

Oligosaccharides produced by the action of fungal cellulase on xyloglucans promoted the elongation of etiolated pea (Pisum sativum L.) stem segments in a straight-growth bioassay designed for the determination of auxins. The oligosaccharides were most active at about 1 micromolar. We tested the relative growth-promoting activities of four HPLC-purified oligosaccharides which shared a common glucose₄· xylose₃ (XG7) core. The substituted oligosaccharides XG8 (glucose₄· xylose₃· galactose) and XG9n (glucose₄· xylose₃· galactose₂) were more effective than XG7 itself and XG9 (glucose₄· xylose₃· galactose· fucose). The same oligosaccharides also promoted the degradation, assayed viscometrically, of xyloglucan by an acidic cellulase from bean (Phaseolus vulgaris L.) leaves. The oligosaccharides were highly active at 10⁻⁴ molar, causing up to a fourfold increase in activity, but the effect was still detectable at 1 micromolar. Those oligosaccharides (XG8 and XG9n) which best promoted growth, stimulated cellulase activity to the greatest extent. The oligosaccharides did not stimulate the action of the cellulase in an assay based on the conversion of [³H]xyloglucan to ethanol-soluble fragments. This suggest that the oligosaccharides enhanced the midchain hydrolysis of xyloglucan molecules (which would rapidly reduce the viscosity of the solution), at the expense of cleavage near the termini (which would yield ethanol-soluble products). We suggest that the promotion of midchain xyloglucan cleavage, by loosening the primary cell wall matrix, explains the promotion of growth by the oligosaccharides.     

Effects of High-voltage Electric Field Treatment on the Water Activity of Bread

Treating bread dough by a high-voltage electric field (HVEF) during the first fermentation enabled the bread dough to retain water in the gluten fibers. The microstructure of the gluten fibers was still visible even with an HVEF treatment at 50 kV for 20 min, but could no longer be observed when the gluten was treated for more than 30 min. The crumb temperature of the HVEF treated bread during baking began to rise a few minutes sooner than that of the untreated bread, but the maximum temperature reached was the same in both cases: 99°C. The fact that the water activity of the HVEF-treated bread was 0.987 ± 0.0056, being higher than that of the untreated bread by 0.011, is in good agreement with the growing tests of Rhizopus nigricans. Furthermore, a distinct decrease in the water loss of the baked gluten was observed after the HVEF treatment. These results suggest that the HVEF treatment increased the ability of the gluten fibers, rather than the starch granules, to absorb and retain water; the state of the remaining water is considered to have become immobile, so that migration of moisture to the starch granules in the bread could be minimized.     

土壤紧实度变化对小麦籽粒产量和品质的影响

以济南17(强筋品种)、烟农15(中筋品种)、鲁麦22(弱筋品种)为供试品种,设置人为碾压和不碾压2种处理,研究了土壤紧实度(以土壤容重表示)变化对不同类型小麦品种的籽粒产量和加工品质的影响。结果表明,随着土壤紧实度提高,3个品种的分蘖成穗率均显著降低,从而导致单位面积穗数和籽粒产量降低。3个品种相比较分蘖成穗率低的鲁麦22籽粒产量降幅最大。相关品质指标测定结果显示,提高土壤紧实度,对3个品种的蛋白质含量、湿面筋含量、沉淀值和吸水率均无显著影响,但济南17的面筋指数明显降低,面团断裂时间和面团稳定时间显著缩短,单位重量面粉烘焙所得的面包体积变小,而烟农15和鲁麦22受影响较小。其原因可能与土壤紧实度提高条件下济南17籽粒中谷蛋白／醇溶蛋白比例和谷蛋白大聚体含量降低有关。将济南17面团流变学特性年际间变化幅度与紧实度变化的处理效应相比较发现,土壤紧实度是影响强筋小麦品种品质性状稳定性的重要因素之一。     

Physicochemical properties of proteins from wheat grown under high-contrast climatic conditions

Studies using electrophoresis, gel chromatography, viscometry, and calorimetry revealed an interrelation of several physicochemical properties of proteins of soft wheat grown under conditions of cool and wet weather with rheological characteristics of gluten and dough and bread quality. The ratio of gliadin and albumin-globulin polypeptides in flour with short-tearing gluten was much lower compared to that in flour with normal gluten. Proteins from flour with short-tearing gluten, including the water-soluble and salt-soluble fraction, had a loose spatial structure. Gluten fractions of this gluten (gliadin and glutenin) were characterized by a more compact and elongated structure compared to normal gluten. As distinct from normal gluten, the conformation of protein particles in short-tearing gluten depended little on hydrophobic interactions. The results suggest that the main components of grain determine the rheological properties of short-tearing gluten.     

Physicochemical properties of proteins from wheat grown under high-contrast climatic conditions

Studies using electrophoresis, gel chromatography, viscometry, and calorimetry revealed an interrelation of several physicochemical properties of proteins of soft wheat grown under conditions of cool and wet weather with rheological characteristics of gluten and dough and bread quality. The ratio of gliadin and albumin-globulin polypeptides in flour with short-tearing gluten was much lower compared to that in flour with normal gluten. Proteins from flour with short-tearing gluten, including the water-soluble and salt-soluble fraction, had a loose spatial structure. Gluten fractions of this gluten (gliadin and glutenin) were characterized by a more compact and elongated structure compared to normal gluten. As distinct from normal gluten, the conformation of protein particles in short-tearing gluten depended little on hydrophobic interactions. The results suggest that the main components of grain determine the rheological properties of short-tearing gluten.     

Cloning, expression and characterization of a family-74 xyloglucanase from Thermobifida fusca.

Thermobifida fusca xyloglucan-specific endo-beta-1,4-glucanase (Xeg)74 and the Xeg74 catalytic domain (CD) were cloned, expressed in Escherichia coli, purified and characterized. This enzyme has a glycohydrolase family-74 CD that is a specific xyloglucanase followed by a family-2 carbohydrate binding module at the C terminus. The Michaelis constant (Km) and maximal rate (Vmax) values for hydrolysis of tamarind seed xyloglucan (tamXG) are 2.4 micro m and 966 micro mol xyloglucan oligosaccharides (XGOs) min-1. micro mol protein-1. More than 75% of the activity was retained after a 16-h incubation at temperatures up to 60 degrees C. The enzyme was most active at pH 6.0-9.4. NMR analysis showed that its catalytic mechanism is inverting. The oligosaccharide products from hydrolysis of tamXG were determined by MS analysis. Cel9B, an active carboxymethylcellulose (CMC)ase from T. fusca, was also found to have activity on xyloglucan (XG) at 49 micro mol.min-1. micro mol protein-1, but it could not hydrolyze XG units containing galactose. An XG/cellulose composite was prepared by growing Gluconacetobacterxylinus on glucose with tamXG in the medium. Although a mixture of purified cellulases was unable to degrade this material, the composite material was fully hydrolyzed when Xeg74 was added. T. fusca was not able to grow on tamXG, but Xeg74 was found in the culture supernatant at the same level as was found in cultures grown on Solka Floc. The function of this enzyme appears to be to break down the XG surrounding cellulose fibrils found in biomass so that T. fusca can utilize the cellulose as a carbon source.     

Genetic analysis of bread-making quality scores in bread wheat using a recombinant inbred line population

Bread-making quality has been evaluated in a progeny of 194 recombinant inbred lines (RILs) from the cross between the two French cultivars Récital and Renan, cultivated in three environments. These cultivars have been previously identified as having contrasting grain protein content and dough rheology properties, although they achieve similar scores for the official bread-making test used for cultivar registration in France. However the progeny displayed a wide range of variations, suggesting that favourable alleles at several loci are present in the two parental lines. Correlation analyses revealed that bread-making scores are poorly correlated among environments, as they are poorly predicted by multiple regression on dough rheology parameters and flour-protein content. However, loaf volume was the most heritable and predictable trait. A total of seven QTLs were found for bread scores, each explaining 5.9–14.6% of trait variation and six for the loaf volume (10.7–17.2%). Most bread-making QTLs, and particularly those detected in all environments, co-located with QTLs for dough rheology, protein content or flour viscosity due to soluble pentosans (Fincher and Stone 1986; Anderson et al. in J Cereal Sci 19:77–82, 1994). Some QTL regions such as those on chromosome 3A and chromosome 7A, which display stable QTLs for bread-making scores and loaf volume, were not previously known to host obvious genes for grain quality.     

Immunogold localization of the cell-wall-matrix polysaccharides rhamnogalacturonan I and xyloglucan during cell expansion and cytokinesis inTrifolium pratense L.; implication for secretory pathways

We have localized two cell-wall-matrix polysaccharides, the main pectic polysaccharide, rhamnogalacturonan I (RG-I), and the hemicellulose, xyloglucan (XG), in root-tip and leaf tissues of red clover (Trifolium pratense L.) using immunoelectron microscopy. Our micrographs show that in both leaf and root tissues RG-I is restricted to the middle lamella, with 80–90% of the label associated with the expanded regions of the middle lamella at the corner junctions between cells. Xyloglucan, however, is nearly exclusively located in the cellulose-microfibril-containing region of the cell wall. Thus, these cell-wall-matrix polysaccharides are present in distinct and complementary regions of the cell wall. Our results further show that during cell expansion both RG-I and XG are present within Golgi cisternae and vesicles, thus confirming that the Golgi apparatus is the main site of synthesis of the non-cellulosic cell-wall polysaccharides. No label is seen over the endoplasmic reticulum, indicating that synthesis of these complex polysaccharides is restricted to the Golgi. The distribution of RG-I and XG in root-tip cells undergoing cell division was also examined, and it was found that while XG is present in the Golgi stacks and cell plate during cytokinesis, RG-I is virtually absent from the forming cell plate.Abbreviations ER endoplasmic reticulum - RG-I rhamnogalacturonan I - XG xyloglucan     

Use of immobilized cell biocatalysts in baking

Baking using baker's yeast immobilized in a starch–gluten–milk matrix (traditional fermented cereal food trahanas), containing viable lactic acid bacteria (LAB), and kefir (natural co-culture of yeasts and LAB) immobilized on orange peel, were investigated. The use of immobilized cells increased shelf life, delayed staling, and improved overall the quality of bread, compared with the traditional baker's yeast bread. These improvements were attributed to the reduction of pH, the lower moisture loss rates, and the presence of LAB, which are known to exhibit antimould properties. Better results were obtained using the sourdough method compared to the straight dough bread-making method. Headspace SPME GC–MS analysis showed that the use of immobilized cells increased the number of bread aroma volatiles, especially esters. The best results, including shelf life and overall bread quality, were obtained in the case of baker's yeast immobilized on trahanas, although kefir immobilized on orange peel seems to be a more cost effective biocatalyst.     

Top-down grafting of xyloglucan to gold monitored by QCM-D and AFM: enzymatic activity and interactions with cellulose

This study focuses on the manufacture and characterization of model surfaces consisting of end-grafted xyloglucan (XG), a naturally occurring polysaccharide, onto a gold substrate. The now well-established XET-technology was utilized for enzymatic incorporation of a thiol moiety at one end of the xyloglucan backbone. This functionalized macromolecule was subsequently top-down grafted to gold, forming a thiol-bonded xyloglucan brushlike layer. The grafting was monitored in situ with QCM-D, and a significant difference in the adsorbed/grafted amount between unmodified xyloglucan and the thiol-functionalized polymer was observed. The grafted surface was demonstrated to be accessible to enzyme digestion using the plant endo-xyloglucanase TmNXG1. The nanotribological properties toward cellulose of the untreated crystal, brush-modified surface, and enzyme-exposed surfaces were compared with a view to understanding the role of xyloglucan in friction reduction. Friction coefficients obtained by the AFM colloidal probe technique using a cellulose functionalized probe on the xyloglucan brush showed an increase of a factor of 2 after the enzyme digestion, and this result is interpreted in terms of surface roughness. Finally, the brush is shown to exhibit binding to cellulose despite its highly oriented nature.     

Relationship between the dough quality and content of specific glutenin proteins in wheat mill streams, and its application to making flour suitable for instant Chinese noodles

The content of specific proteins such as high-molecular-weight glutenin subunits HMW-GS 5+10 and low-molecular-weight glutenin subunits LMW-GS KS2 in wheat mill streams of extra-strong Kachikei 33 wheat was quantified by SDS-PAGE and 2D-PAGE. The mill streams showed varied quantities of HMW-GS 5+10 (0.077 to 2.007 mg/g of mill stream), LMW-GS KS2 (0.018 to 0.586 mg/g of mill stream) and total protein (9.42% to 18.98%). The contents of these specific proteins in the mill streams were significantly correlated with the SDS sedimentation volume and the mixing properties, which are respective indices of specific loaf volume and dough strength. The contents of these specific glutenin proteins in the mill streams were therefore found to be significantly important for improving the dough quality suitable for bread and Chinese noodles. Accordingly, we present here the application of this information to the development of an effective method for producing mill streams with high quality and yield that are suitable for instant Chinese noodles.     

Behavior of 2-O-α-D-Glucopyranosyl-L-ascorbic Acid in a Flour-water Suspension and Its Effectiveness as a Dough Improver

The behavior of 2-O-α-D-glucopyranosyl-L-ascorbic acid (AA-2G) in a whole wheat flour suspension was investigated. AA-2G was hydrolyzed by a non-dialyzable and heat-labile component in flour and liberated L-ascorbic acid (AA). The pH profile for the hydrolysis is similar to that of rice α-glucosidase. However, the hydrolysis of AA-2G was inhibited completely by endogenous sugars, mainly maltose, which were produced rapidly during the hydration of flours. In the presence of Saccharomyces cerevisiae strain with a strong maltose-utilizing capability, the hydrolysis of AA-2G proceeded in a flour suspension, and was followed by the formation of dehydro-L-ascorbic acid. The hydrolysis of AA-2G also proceeded in yeasted dough, concomitant with increases in the resistance on an extensigram and in the loaf volume of the bread. These effects of AA-2G on dough were less than those of equimolar AA because of the imperfect liberation of AA. The results show that AA-2G could be useful as a highly stable dough improver.