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1.
A procedure was established for isolating simultaneously high molecular weight (HMW-GS) and low molecular weight glutenin subunits (LMW-GS) from chloroform-defatted wheat flour. Temperature conditions were optimised to obtain maximum yield and purity. A pre-extraction of gliadin was performed with 50% (v/v) propan-1-ol at room temperature. Glutenin subunits were solubilised from the residue at 60 °C with 50% (v/v) propan-1-ol containing 1% (w/v) dithiothreitol. The HMW-GS in the glutenin mixture were then precipitated selectively by increasing the propan-1-ol concentration to 60%. Upon addition of propan-1-ol to a final concentration of 85%, LMW-GS were precipitated. It was shown by densitometric scanning of SDS–PAGE gels that the isolated glutenin subunits were very pure. Furthermore, a partial separation of LMW-GS into B and C type subunits was observed when the propan-1-ol concentration was increased gradually.  相似文献   

2.
Both genetic and environmental factors influence the types and amounts of wheat proteins that link together to form polymers essential for flour quality. To understand how plant growth conditions might influence gluten polymer formation, protein fractions containing small and large polymers were separated from flour from the US wheat Butte 86 grown in the absence or presence of post-anthesis fertilizer. Proteins in the polymer fractions were analyzed by quantitative two-dimensional gel electrophoresis (2-DE). The ratio of high molecular weight glutenin subunits (HMW-GS) to low molecular weight glutenin subunits (LMW-GS) increased in both fractions in response to fertilizer, due in part to small increases in the proportions of individual HMW-GS. There were also changes within the LMW-GS. In particular, omega and alpha chain terminators increased in proportion in both polymer fractions, but changes were more pronounced in the large polymer fractions. Serpins also increased in both polymer fractions. Additionally, the study revealed differences in the proportions of traditional LMW-GS in small and large polymer fractions. LMW-s type proteins were more abundant in the large polymers while LMW-i type proteins were more prevalent in the small polymers, suggesting that these proteins may play different roles in the gluten polymer.  相似文献   

3.
The breadmaking quality of wheat is affected by the composition of gluten proteins and the polymerisation of subunits that are synthesised and accumulated in developing wheat grain. The biological mechanisms and time course of these events during grain development are documented, but not widely confirmed. Therefore, the aim of this study was to monitor the accumulation of gluten protein subunits and the size distribution of protein aggregates during grain development. The effect of desiccation on the polymerisation of gluten proteins and the functional properties of gluten were also studied. The results showed that the size of glutenin polymers remained consistently low until yellow ripeness (YR), while it increased during grain desiccation after YR. Hence, this polymerisation process was presumed to be initiated by desiccation. A similar polymerisation event was also observed when premature grains were dried artificially. The composition of gluten proteins, the ratios of glutenin to gliadin and high molecular weight-glutenin subunits to low molecular weight-glutenin subunits, in premature grain after artificial desiccation showed close association with the size of glutenin polymers in artificially dried grain. Functional properties of gluten in these samples were also associated with polymer size after artificial desiccation.  相似文献   

4.
D-type low molecular weight subunits of bread wheat glutenin have ω-gliadin type N-terminal amino acid sequences, but are incorporated into the glutenin polymers because of the presence of cysteine residues. In order to determine the number and position of cysteine residues, ID-encoded D-type low molecular weight subunits of wheat glutenin were purified from the cv. Chinese Spring using a procedure that allowed high recovery. Comparison of the molecular weights of alkylated and unalkylated subunits by MALDI mass spectrometry indicated the presence of only one cysteine residue per molecule. This was supported also by the detection of dimers of D subunits in gluten. An internal sequence of 62 amino acids preceding the cysteine was obtained, but it was not possible to identify the cysteine residue, either because it was not within the range of N-terminal sequencing of peptides obtained, or because it was present in one of the two unidentified positions.  相似文献   

5.
Low molecular weight (LMW-GS) and high molecular weight glutenin subunits (HMW-GS) were added to a base flour using both «addition» and «incorporation» protocols. «Incorporation» of glutenin subunits into the glutenin network of the base flour was performed by partial (reversible) reduction and subsequent reoxidation of the glutenin network in the presence of the added glutenin subunits whereas, in the «addition» protocol, glutenin subunits were added without reduction/oxidation. The effects of both «addition» and «incorporation» of alkylated and unalkylated LMW-GS and HMW-GS on dough extension parameters maximum resistance (MR) and extensibility (EX) were compared and thoroughly discussed. HMW-GS and LMW-GS had totally different effects on dough extensibility. «Addition» of LMW-GS significantly decreased both MR and EX whereas HMW-GS caused a significant increase in MR. «Incorporation» of LMW-GS caused a decrease in MR whereas HMW-GS clearly increased MR. The similarity in effects obtained with «addition» and «incorporation» of glutenin subunits indicated that, even with «addition», glutenin subunits can be partially incorporated into the glutenin network in the presence of oxygen. Alkylated and unalkylated glutenin subunits had different effects. This was probably caused by the effect of free sulphydryl groups in unalkylated subunits (possibility of SS/SH exchanges and/or incorporation) and/or the effect induced by introduction of alkylated derived substituents. A protocol for «incorporation with excess KIO3» was developed to exclude the possible effect of a lowering of the available oxidant concentration by oxidation of free sulphydryl groups in glutenin subunits. However, the use of high levels of oxidant in the «incorporation with excess KIO3» protocol seems to overrule the effects of added glutenin subunits or may force glutenin subunits to incorporate differently from what can be observed under gentle oxidation conditions. Therefore, «incorporation with excess KIO3» is not suitable for studying the effects of incorporation of glutenin subunits on dough extensibility.  相似文献   

6.
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.  相似文献   

7.
Glutenin polymers are formed by high (HMW-GS) and low molecular weight glutenin subunits (LMW-GS). The latter group of subunits has been less characterised compared to the former due to their great number and heterogeneity.  相似文献   

8.
小麦贮藏蛋白特性及其遗传转化   总被引：13，自引：7，他引：13  
小麦籽粒贮藏蛋白由醇溶蛋白和谷蛋白组成。醇溶蛋白在组成上以单体形式存在 ,具有高度的异质性和复杂性。它决定小麦面筋的粘性。谷蛋白是由多个亚基组成的高分子聚合体 ,决定面筋的弹性。它可分为低分子量谷蛋白亚基和高分子量谷蛋白亚基 (HMW- GS)。HMW- GS具有相似的分子结构 ,即由中央重复序列、无重复的 N端和 C端组成。HMW- GS对小麦烘烤品质起着决定性作用 ,但因 HMW- GS类型不同而对加工品质的贡献大小各异。许多 HMW- GS基因已被揭示。实践证明 ,利用基因枪法 ,将 HMW- GS基因导入普通小麦的细胞核内 ,能够达到改良小麦烘焙品质的目的。随着分子生物学技术的不断发展 ,可望从营养和加工角度来改良小麦品质的特性  相似文献   

9.
The low molecular weight glutenin subunits (LMW-GS) are wheat storage proteins participating to the formation of glutenin polymers that, along with the other gluten proteins, allow the accumulation of a large quantity of protein in the endosperm tissue. The size and composition of the glutenin polymers are directly related to gluten visco-elastic properties. In particular, LMW-GS composition is the factor most influencing durum wheat quality.  相似文献   

10.
分子生物学技术在普通小麦谷蛋白研究中的应用   总被引：8，自引：3，他引：8  
小麦谷蛋白是面筋的主要成分之一，对小麦食品的加工品质起着重要作用。本文从基因序列、分子结构、多态性、遗传转化、QTL研究和MAS等方面综述了国内外有关麦谷蛋白亚基的研究进展。这些研究结果表明，麦谷蛋白的等位基因变异十分丰富，多态性高．序列之间存在很高的同源性。麦谷蛋白的基因结构分为三部分：无重复结构的N-末端和C-末端以及中部重复区域，等位基因的变异主要由基因中部重复区域的序列大小、重复次数及该区域内DNA序列的插入或缺失所造成；Cys-残基的数目和位置影响麦谷蛋白聚合体内亚基间的相互作用，是影响亚基生化特性的重要因素；应用转基因技术已将HMW-GS基因(1Ax1、1Dx5和1Dy10)导入普通小麦中，有助于进行品质改良和麦谷蛋白结构与功能的深入研究。此外，对面筋强度性状的QTL分析和分子标记辅助育种也进行了阐述。  相似文献   

11.
小麦低分子量谷蛋白一级结构分析   总被引：1，自引：1，他引：1  
用数学统计方法和蛋白质序列分析软件对部分已测序的17种低分子量走谷蛋白亚基含氮量、氨基酸含量及一级结构序列进行比较分析。以期从氨基酸层次上揭示其结构和功能及进化上的关系．结果表明：(1)其平均含氮量为17．8％;(2)氨基酸含量基本恒定。以谷氨酰胺的含量最多。且常连接在一起;(3)存在119个保守性氨基酸残基扣两个保守性片段。一个是“QQQ-PPFSQQ”。一个是“IP-VHPSILQQLNPCKVFLQQ—C—P—AM-Q-LARSQM-QS-CHVMQQQCCQQL—QIPQQSRYEAI-AI-YSIILQEQQ”(“-”表示各序列中不同的氨基酸);(4)X84960、X84961、Y14104;X13306、U86025、U86027、U86029两组亚基序列中各具有一定程度的同源性。  相似文献   

12.
Molecular weight and size distributions of two glutenin polymers were determined by a multi-angle laser light scattering (MALLS) photometer on-line to a size exclusion chromatography (SEC) system. Two glutenin polymers extracted, sonicated and purified from wheat flours of different cultivars, i.e. Cheyenne and Chinese Spring, were accurately fractionated by SEC using three buffers (pH 2.6, 4.0 and 6.9) and two column sets. Both molecular weight distribution (MWD) and radius of gyration distribution (RGD) could be used to differentiate the two cultivars. MWD of glutenin polymers is a complex mixture of high- and low-molecular weight fractions and the relative percentage was found to be very different. The two cultivars were found to be different; in particular, the Chinese Spring polymer showed more compact conformation than the Cheyenne polymer. The slope of the conformation plot for glutenin was about 0.37, close to the theoretical value for compact spheres. Determination of glutenin MWD and RGD was difficult and depended on the buffer used and the SEC columns.  相似文献   

13.
The variations of the amounts of individual high molecular weight glutenin subunits (HMW-GS), of the ratios HMW-GSy to HMW-GSx and HMW-GS to low molecular weight glutenin subunits (LMW-GS) and of protein content were evaluated for eight durum wheat cultivars in two regions using four fertilizer combinations during two successive years. All measured parameters showed significant variation with genotypes (G), environments (E) and fertilizers (F). The interaction E × G × F was highly significant for glutenin amount variation. Amongst cultivars possessing HMW-GS 20, landraces seem to better value the N-fertilizer use for the accumulation of HMW-GSy than high yielding cultivars. Both HMW-GSy to HMW-GSx and HMW-GS to LMW-GS ratios were found to be positively correlated (p < 0.05) with total protein content.  相似文献   

14.
小麦谷蛋白大聚合体含量的影响因素   总被引：18，自引：0，他引：18  
小麦籽籽谷蛋白大聚合体（ＧＭＰ）的含量反映了谷蛋白聚合体的粒度分布情况,其高低与其他烘烤品质性状有密切的关系。本文对其影响因素进行了研究。结果表明,ＧＭＰ含量受Ｇｌｕ－１位点等位基因变异、亚基相对含量和环境因素等的影响。其中,Ｇｌｕ－Ａ１ｂ和Ｇｌｕ－Ｄ１ｄ对ＧＭＰ含量有正效应,而Ｇｌｕ－Ａｌｃ和Ｇｌｕ－Ｄ１ａ对ＧＭＰ含量有负效应;Ａ组和Ｃ组亚基的相对含量分别与ＧＭＰ含量正相关和负相关。ＧＭＰ含量也  相似文献   

15.
The sample preparation method of high molecular weight glutenin subunits (HMW-GS) for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis, without a separation step, by high-performance liquid chromatography (HPLC) was established in this study. Three major factors influencing mass spectra—the ratio of components of the solvent, the resolving time, and the sample volume—were optimized using HMW-GS mixtures extracted from Chinese cultivar Jing 411. The results showed that the optimized method for sample preparation was to resolve HMW-GS from 20 mg in an hour with 50 μl solvent of 0.4% TFA, 30.0% ACN and 69.6% H2O. The stable mass spectra and accurate molecular weights of 16 major HMW glutenin subunits from common wheat and related species were obtained using the optimized MALDI-TOF-MS method. Seven subunits, where each was from 2–5 cultivars, showed very similar molecular weights. The determined molecular weights of 11 subunits were close to those calculated from their coding sequences. In addition, no positive reaction between HMW-GS and GelCode® Glycoprotein Staining Reagent was observed. These results suggested that HMW-GS lack extensive post-translational modifications (PTMs), but low levels of glycosylation or phosphorylation present in some subunits cannot be ruled out. Because of its ability to obtain a rapid, complete and precise profile of HMW glutenin subunits without purifying procedures, MALDI-TOF-MS is expected to be a powerful technique for structural and functional studies of HMW glutenin subunits as well as other cereal proteins.  相似文献   

16.
甘肃小麦品种(系)HMW-GS遗传变异分析   总被引：4，自引：1，他引：4  
为了了解甘肃省小麦品种HMW-GS的遗传变异和组成，为甘肃优质专用小麦品种筛选和品种改良提供依据,采用SDS-PAGE法，分析了254份甘肃省小麦材料(育成品种(系)和农家品种)Glu-1位点的HMW-GS变异，共检测到22种HMW-GS变异，Glu-A1位点3种，Glu-B1位点11种，Glu-D1位点8种。其中110个育成品种(系)的15种HMW-GS有27种亚基组合类型。Glu-A1位点有2种亚基，47．3％的品种该位点具有优质亚基；Glu-B1位点有7种亚基，76．4％的品种具优质亚基；Glu-D1位点有6种亚基，32．7％的品种具优质亚基。14．5％的品种(n：15)Glu-1位点具优质亚基。144份农家品种的18种HMW-GS共有29种亚基组合形式，Glu-A1位点有3种亚基,Glu-B1位点有9种亚基，Glu-D1位点有6种亚基，无优质亚基组合。  相似文献   

17.
This study aims to understand the environmental factors, focusing on rain and fungal infection, affecting the assembly of glutenin polymers during grain maturation. Spring wheat was grown in the field and grains were sampled from 50% grain moisture until maturity. Grain moisture content, protein content, size of glutenin polymers, the presence of proteases, and the amount of DNA from common wheat pathogenic fungi were analysed.Rain influenced the rate of grain desiccation that occurred parallel to the rate of glutenin polymer assembly. Rapid desiccation contributed to faster glutenin polymer assembly than gradual desiccation. Severe reduction in the glutenin polymer size coincided with increased grain moisture due to rain. Furthermore, increased fungal DNA followed by presence of gluten-degrading proteases was observed in the grain after humid conditions. The presence of gluten-degrading proteases was presumably involved in reducing the size of glutenin polymers in grain.Our study gave new insight into how environmental conditions could be associated with the assembly of glutenin polymers during grain maturation. The results suggest that rain and/or fungal proteases play an important role in reducing the molecular size of glutenin polymers.  相似文献   

18.
The effects of 60Co gamma-irradiation treatments (2·5, 5·0, 10·0 and 20·0 kGy) on the gluten proteins of two bread wheats and one durum wheat cultivar were investigated. Dough rheological properties of the flour processed from the irradiated wheat were also determined using a computerised micromixograph. Irradiation caused a significant deteriorating effect on all mixogram parameters. There was no observable effect of irradiation on gliadin proteins analysed by polyacrylamide gel electrophoresis. The 50% 1-propanol-insoluble (50 PI) glutenin fraction was highly affected by irradiation. By sodium dodecyl sulfate polyacrylamide gel electrophoresis, reduced 50 PI glutenin showed a noticeable reduction in band intensities of both high (HMW) and low molecular weight (LMW) glutenin subunits (GS) with increasing irradiation dosage greater than 5 kGy. The irradiation effect on 50 PI glutenin was further studied and quantified by reversed-phase high-performance liquid chromatography of glutenin subunits; there was a progressive decrease in the quantity of subunits with increasing irradiation dose level. Compared to non-irradiated wheat, the relative decline in total insoluble glutenin at the 20 kGy dosage level ranged from 34–49% depending on cultivar. Increasing levels of irradiation also progressively reduced the ratio of HMW:LMW-GS up to 13–15% at 20 kGy indicating that irradiation had a greater effect on the largest polymers of glutenin. The observed weakening of dough mixing properties and concomitant decline in the quantity of 50 PI glutenin with increasing levels of gamma-irradiation are consistent with a degradation of glutenin to a lower average molecular size by depolymerisation and/or disaggregation.  相似文献   

19.
To understand more precisely the function of free glutenin SH and SS groups in glutenins of developing wheat for UPP formation, a specific sulfhydryl probe, monobromobimane (mBBr), was used for an in vitro protein labeling. By applying this procedure to two varieties of wheat differing in high molecular weight glutenin subunit composition (2*, 7+8, 5+10 and 0, 6+8, 2+12, respectively, for Soissons and Thésée), we showed that the major wheat glutenin subunits residing in the protein body undergo redox change during the development and the maturation of the grain. Indeed, during the cell division and the cell enlargement phase, glutenin subunits and particularly LMW-GS have a large amount of free SH groups and become oxidized during grain dehydration which coincided with the formation of UPP. Moreover, mBBr derivatization of free glutenin SH groups before the artificial grain desiccation inhibits the UPP deposition. As HPSEC-MALLS analysis showed, the alkylation of free glutenin SH groups before the desiccation induces an increase in the SDS solubility of the polymeric proteins by reducing both their molecular weight distribution and their compactness. These results are discussed in connection with an ‘hyperaggregation model’ which has been proposed to explain the formation of glutenin polymer in wheat kernel.  相似文献   

20.
为从一级结构水平揭示小麦高分子量麦谷蛋白亚基的结构、功能及其进化上的关系,利用生物信息学软件(DNAMAN)对已在GenBank数据库中登录的22个小麦高分子量麦谷蛋白亚基序列进行了分析.结果表明,这些高分子量谷蛋白亚基平均含氮量为16.186%;其各种氨基酸含量基本恒定,其中谷氨酰胺含量最高(33.74%),且主要以QQ形式连接在一起,这种结构特点可能影响小麦面粉的加工品质;同时在一级结构序列上还发现有232个保守的氨基酸位点(56个单个氨基酸座位和一些长短不同的氨基酸片段),较大的保守性氨基酸片段主要为AEGEAS-QLQCER/HEL,GSFY PG/SETTP-QQLQQ,YYPGQAS/FP/SQQ/RP/SGQG/RQQ,PGQGQQ/PG/A/YYPTS-QQ等;分别有6组共13个序列相似度均达到100%,推测这些高分子量谷蛋白亚基在进化上具有较强的保守性.  相似文献   

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