红苋R104种子谷蛋白亚基组成及二硫键连接蛋白研究

应用还原和非还原单向ＳＤＳ－ＰＡＧＥ及二者结合的双向电泳技术，研究了红苋Ｒ１０４种子谷蛋白的亚基组成。谷蛋白由多种亚基组成，高分子量二硫键连接蛋白占其多数，经还原裂解成Ａ（５４ＫＤ）、Ｂ（４０ＫＤ，３９ＫＤ）、Ｃ（３３ＫＤ，３１ＫＤ）、Ｄ（２２ＫＤ，２０ＫＤ，１８ＫＤ，）和Ｅ（１５ＫＤ）５组主要单肽链单元，提出了二硫键连接蛋白的组成模式，并发现低分子量谷蛋白亚基存在肽内二硫键     

Disulphide structure of high-molecular-weight (HMW-) gliadins as affected by terminators

This study aimed at elucidating SS-bonds of HMW-gliadins (HGL) from wheat with the focus on terminators of glutenin polymerisation. HGL from wheat flour extracts non-treated or treated with the S-alkylation reagent N-ethylmaleinimide (NEMI) were compared. HGL from wheat flour Akteur were isolated, hydrolysed with thermolysin and the resulting peptides pre-separated by gel permeation chromatography and analysed by liquid chromatography/mass-spectrometry using alternating electron transfer dissociation/collision-induced dissociation. Altogether, 22 and 28 SS-peptides from samples without and with NEMI treatment, respectively, were identified. Twenty-six peptides included standard SS-bonds of α- and γ-gliadins, high-molecular-weight and low-molecular-weight glutenin subunits. Eleven SS-bonds were identified for the first time. Fifteen peptides unique to HGL contained cysteine residues from gliadins with an odd number of cysteines (ω5-, α- and γ-gliadins). Thus, gliadins with an odd number of cysteines, glutathione and cysteine had acted as terminators of glutenin polymerisation. Decisive differences between samples without and with NEMI treatment were not obvious showing that the termination of polymerisation was already completed in the flour. The two HGL samples, however, were different in the majority of ten peptides that included disulphide-linked low-molecular-weight (LMW) thiols such as glutathione and cysteine with the former being enriched in the non-treated HGL-sample.     

Identification of a Group of Novel y-Gliadin Genes

γ-Gliadins are an important component of wheat seed storage proteins. Four novel γ-gliadin genes （Gli-ngl to Gli-ng4） were cloned from wheat （Triticum aestivum） and Aegilops species. The novel γ-gliadins were much smaller in molecular size when compared to the typical γ-gliadins, which was caused by deletion of the non-repetitive domain, glutamine-rich region, 3＂ part of the repetitive domain, and 5＇ part of the C-terminal, possibly due to illegitimate recombination between the repetitive domain and the C-terminal. As a result, Gli-ngl and Gli-ng4 only contained two and three cysteine residues, respectively. Gli-ngl, as the representative of novel γ-gliadin genes, has been sub-cloned into an Escherichia coli expression system. SDS- PAGE indicated that the both cysteine residues of Gli-ngl could participate in the formation of intermolecular disulphide bonds in vitro. Successful cloning of Gli-ngl from seed cDNA of T. aestivum cv. Chinese Spring suggested that these novel γ-gliadin genes were normally transcribed during the development of seeds. Phylogenic analysis indicated that the four novel γ-gliadin genes had a closer relationship with those from the B （S） genome of wheat.     

Purification and Characterisation of HighMrGlutenin Subunit 20 and its Linked y-type Subunit from Durum Wheat

HighM_rglutenin subunit 20 and its linked y-type subunit, present in the durum wheat cultivar Lira, were purified by preparative reversed-phase high-performance liquid chromatography (RP–HPLC). Amino acid and N-terminal sequence analysis of subunit 20y confirmed that it corresponded to a y-type subunit. Moreover, the number and position of the cysteine residues in subunit 20 were determined by alkylation with the fluorogenic reagent 7-fluoro-4-sulfamoyl-2,1,3,-benzoxadiazole (ABD-F) and subsequent enzymic digestion with trypsin. N-terminal amino acid sequence analysis of the fluorescent peptides showed that subunit 20 had only two cysteine residues, one in the N-terminal region and the other in the C-terminal domain.     

Chemical Soil Fumigation for Strawberry Nursery in Jalisco (Mexico): 2013–2014 Results of Semarnat/Unido Project

ABSTRACT

We evaluated the effectiveness of alternative fumigants on weed control and plant productivity in two nurseries located at high elevation and low latitude (Ciudad Guzman, Jalisco), during 2013 and 2014 at Driscoll’s and Lassen Canyon South nurseries. Treatments were: MB:chloropicrin (MB:CP); 1,3-dichloropropene:CP; CP alone; metam sodium (MS) alone; dimethyl disulphide (DMDS):CP; and sequentially applied CP and MS. A rotary spading machine was used for MS and CP followed by MS. All treatments produced similar pest suppression and runner-plant yields during 2013 and 2014. However, CP alone showed very poor control of the most abundant weed (barnyardgrass, Echinochloa crus-galli). Two years of work on MB alternatives were not sufficient to provide reliable recommendations on this critical need, therefore, the Methyl Bromide Technical Options Committee recommended critical use nomination for MB in 2015.     

Disulphide Bonds in Wheat Gluten Proteins

Disulphide bonds play a key role in determining the structure and properties of wheat gluten proteins. Comparison of the sequences of monomeric gliadins and polymeric glutenin subunits allows the identification of conserved and variant cysteine residues. Direct disulphide bond determination demonstrates that the conserved cysteine residues present in S-rich prolamins (α-type gliadins, γ-type gliadins and LMW subunits) form intra-chain disulphide bonds while additional cysteines residues present only in the LMW subunits form inter-chain bonds with cysteines in HMW subunits and other LMW subunits. Conserved and variant cysteine residues are also present in the HMW subunits but their patterns of disulphide bond formation are less well understood. Further information on the abilities of individual cysteine residues to form intra- and inter-chain disulphide bonds has also been obtained by heterologous expression of wild type and mutant proteins inE. coliand, in the case of the HMW subunits, by examination of the patterns of dimers recovered on partial reduction of glutenin or resulting from the expression of subunits in transgenic tobacco plants. Wheat gluten proteins are folded and assembled within the lumen of the endoplasmic reticulum of the developing endosperm cells, where disulphide bond formation and exchange may be catalysed by the enzyme protein disulphide isomerase. Similarly, disulphide bond reduction, for example to facilitate mobilisation during germination, may be catalysed by thioredoxinh. Understanding the mechanism and specificity of disulphide bond formation in gluten is crucial for the manipulation of its functional properties by genetic engineering or chemical modification.     

Proteins in Rice Grains Influence Cooking Properties!

The Australian rice industry is small compared to others. In order to compete, the industry targets the high-quality end of niche markets. To maintain quality it is important to understand quality attributes and to extract the most information from tests to evaluate quality. Viscosity of the flour (RVA) is a useful tool for evaluating quality. The aim of this study is to determine the effect of proteins on forming a viscosity curve, and then to understand how proteins affect the quality of cooked rice. The results demonstrate clearly that proteins influence viscosity curves both through binding water, which increases the concentration of the dispersed and viscous phase of gelatinised starch and through the agency of a network linked by disulphide bonds. Increasing rates of N nutrition decreases the peak height, consistent with fewer disulphide bonds so less contribution from the linked network. Storage also decreases peak height by increasing disulphide bonds. Proteins affect the amount of water the rice absorbs early in cooking, and the availability of water early in cooking will determine the hydration of the protein and the concentration of the dispersed and viscous phases of the starch, which will determine the texture of the cooked rice.     

Effect of fish feed processing conditions on digestive protease activities, free amino acid pools, feed conversion efficiency and growth in Atlantic salmon (Salmo salar L.)

The responses of the digestive proteases trypsin and chymotrypsin and protein metabolism to differences in feed protein quality were investigated in Atlantic salmon (Salmo salar L.). Two sets of experimental feeds were produced. Each set of high and low quality feeds was provided to either 150 g or 2 kg salmon. Protein in the high quality feeds had significantly higher percentages of free (reactive) sulphydryl (SH) groups than the corresponding feeds based on low quality meals. After 90 days feeding, groups given high and low quality feeds did not differ in their specific growth rates (SGR) in either experiment. However, feed conversion efficiency (FCE) was significantly different between the high and low quality feed groups in 2 kg salmon, where the difference between the high and low feed protein qualities was larger, 10% versus 4% SH/[SH + (S–S)] in 150 g salmon. Higher FCE was preceded by significantly higher trypsin and chymotrypsin specific activities on day 60. SGR, in general, changed after the first month and was stable during the last 2 months in both experiments. Concurrently, both trypsin (T) and chymotrypsin (C) decreased with an increased activity ratio of trypsin to chymotrypsin (T/C ratio), and resulting in significantly lower T/C ratio on day 90 in salmon feeding on high quality feeds in both sizes of fish. Differences in FCE were associated with significant differences in levels of total free amino acids (TFAA) in the plasma and the white muscle, as well as in the ratio of essential to non‐essential free amino acids (EAA/NEAA ratio), free hydroxyproline, and RNA in the white muscle. Interestingly, after 3 days starvation (day 93), 5–7 h postprandial EAA/NEAA ratio in the plasma was significantly lower in the high quality diet groups in both experiments. Trypsin specific activity inversely correlated with muscle TFAA levels in 2 kg salmon, concurrent with higher muscle levels of RNA, lower free hydroxyproline and higher FCE in fish fed higher quality diets.