Characterisation of proteolytic enzymes of Eurygaster integriceps Put. (Sunn bug), a major pest of cereals

Eurygaster integriceps (Sunn pest or Sunn bug) is one of the most significant pests of wheat and is responsible for substantial losses in yield and quality of wheat grain in Europe and Asia. Sunn pest salivary gland-derived proteases and other hydrolases damage grain proteins and starch. Characterisation of protease activities from both Sunn pest salivary glands and Sunn pest-damaged wheat grains revealed a broad range of activities in terms of substrate specificity and diversity of isoelectric point. Neutral and alkaline proteases present in Sunn pest-damaged grains were shown to be capable of hydrolyzing gluten proteins, whilst some proteases were also shown to be active against gelatin. The neutral serine proteases present play the dominant role in degradation of gluten quality. The sensitivity of some proteases to proteinaceous and non-proteinaceous serine proteinase inhibitors was shown, including that of a recombinantly expressed protease. It was found that proteases isolated from Sunn pest salivary glands could be activated by trypsin indicating that they are present as zymogens in vivo. Analysis of individual Sunn pest-damaged grains showed great diversity in the proteases present. This work highlights the challenges of developing proteinase inhibitors to manage Sunn pest damage

Colloidal stability of tannins: astringency, wine tasting and beyond

Tannin-tannin and tannin-protein interactions in water-ethanol solvent mixtures are studied in the context of red wine tasting. While tannin self-aggregation is relevant for visual aspect of wine tasting (limpidity and related colloidal phenomena), tannin affinities for salivary proline-rich proteins is fundamental for a wide spectrum of organoleptic properties related to astringency. Tannin-tannin interactions are analyzed in water-ethanol wine-like solvents and the precipitation map is constructed for a typical grape tannin. The interaction between tannins and human salivary proline-rich proteins (PRP) are investigated in the framework of the shell model for micellization, known for describing tannin-induced aggregation of beta-casein. Tannin-assisted micellization and compaction of proteins observed by SAXS are described quantitatively and discussed in the case of astringency

Antibodies raised against a Sunn bug (Eurygaster integriceps Put.) recombinant protease, rGHP3p2, can inhibit gluten‐hydrolyzing activity

Sunn pest or Sunn bug, Eurygaster integriceps Put., salivary gland proteases are responsible for the deterioration of wheat flour quality during dough mixing, resulting from gluten hydrolysis. These proteases are highly heterogeneous and show low sensitivity to most types of proteinaceous inhibitors, meaning that such inhibitors cannot be used to prevent gluten damage. The present study describes the generation of a specific peptide antibody, raised against the active center of the recombinant gluten-hydrolyzing protease (GHP3). The recombinant protein, encoding two repeats of the GHP3 sequence element involved in forming the S4 pocket and binding of substrate at position P4, was designed and expressed in Escherichia coli. The antibodies raised to this recombinant protein showed inhibitory activity against the GHP3 protease. The results indicate that it is possible to design specific antibodies to inhibit wheat-bug gluten-hydrolyzing proteases

Identification of duplicate accessions in the sweet maize collection by means of zein electrophoresis

Of all the subspecies of Zea mays L. cultivated in the world, sweet maize is the most important for the global economy. The leading seed-growing companies and research institutions around the world are engaged in breeding this crop. To meet the increasing demands of the industry to grain quality, it is important to select appropriate local varieties and lines for hybridization. Local (usually heterogeneous) varieties are a valuable source material for creating self-pollinated lines that contribute to a significant broadening of the genetic base of parental forms used in breeding. The advantages of sweet maize varieties and the interest of the food industry in them make it possible to consider accessions from the maize collection of the N.I. Vavilov Institute (VIR) as a potentially valuable source material for breeding. The present research concentrated on 19 local sweet maize varieties with different grain colors from the VIR collection, that is, 9 varieties with the blue color of ripe grain, 4 with white (colorless) grain, 3 with yellow, and 3 with red. The research included an analysis of zein electrophoretic patterns (protein markers); a study of their biotype composition and the nature of genetic polymorphism, as well as the creation of a protein pattern database for each accession. For a series of accessions with the same varietal name, but different catalog numbers, the degree of their identity was determined from their biotype composition in order to exclude duplication. Zein electrophoresis was carried out in vertical plates of 10 % polyacrylamide gel according to the standard ISTA technique developed with the participation of the Biochemistry and Molecular Biology Department of VIR. Zein patterns were used for the first time to electrophoretically study sweet maize varieties with different grain colors. Unique zein patterns were established for all the accessions studied, which makes possible their identification by specific marker components. The results of this work characterize zein electrophoresis as a useful tool for the identification and registration of duplicate accessions in the VIR collection of sweet maize varieties

Zein patterns as effective markers of valuable agronomic traits in maize

The effective use of maize (Zea mays L.) in agriculture depends entirely on the knowledge of its gene pool, which, in turn, needs to undergo registration and certification as new data become available. Maize is one of the model objects for applying the marker methodology. This article summarizes the results of a long-term study of the maize gene pool at N. I. Vavilov All-Russian Institute of Plant Genetic Resources (VIR) in two main areas: identification of the world genetic diversity of maize (including the most important wild relatives for breeding use), and search for reliable protein (zein) markers of traits valuable for breeding in order to use them in improvement, seed production, and seed control of this crop. Specific examples show the effectiveness of using zein markers in the development of new inbred lines and improvement of the existing ones. On the examples of such agronomic traits as early maturity, explosiveness, and waxiness of maize grain, the possibility of controlling the “inclusion” of these traits in the breeding material with the help of zein markers has been demonstrated. Evidence of the effectiveness of applying zein markers in the selection of parental pairs to obtain highly heterotic hybrids is presented. The nomenclature of zein adopted at VIR after its separation by polyacrylamide gel electrophoresis (PAGE) in an acidic buffer was compared with the nomenclature of zein after polyacrylamide gel electrophoresis with sodium dodecyl sulfate (SDS-PAGE) adopted among the foreign scientific community. The studies were performed on a large volume of accessions from the VIR collection as well as on breeding material provided by the country’s leading breeders

Glutamate 270 plays an essential role in K activation and domain closure of Thermus thermophilus isopropylmalate dehydrogenase

The mutant E270A of Thermus thermophilus 3-isopropylmalate dehydrogenase exhibits largely reduced (∼1%) catalytic activity and negligible activation by K+ compared to the wild-type enzyme. A 3–4 kcal/mol increase in the activation energy of the catalysed reaction upon this mutation could also be predicted by QM/MM calculations. In the X-ray structure of the E270A mutant a water molecule was observed to take the place of K+. SAXS and FRET experiments revealed the essential role of E270 in stabilisation of the active domain-closed conformation of the enzyme. In addition, E270 seems to position K+ into close proximity of the nicotinamide ring of NAD+ and the electron-withdrawing effect of K+ may help to polarise the aromatic ring in order to aid the hydride-transfer

Variation in the organization and subunit composition of the mammalian pyruvate dehydrogenase complex E2/E3BP core assembly

The final version of this article is available at the link below.Crucial to glucose homoeostasis in humans, the hPDC (human pyruvate dehydrogenase complex) is a massive molecular machine comprising multiple copies of three distinct enzymes (E1–E3) and an accessory subunit, E3BP (E3-binding protein). Its icosahedral E2/E3BP 60-meric ‘core’ provides the central structural and mechanistic framework ensuring favourable E1 and E3 positioning and enzyme co-operativity. Current core models indicate either a 48E2+12E3BP or a 40E2+20E3BP subunit composition. In the present study, we demonstrate clear differences in subunit content and organization between the recombinant hPDC core (rhPDC; 40E2+20E3BP), generated under defined conditions where E3BP is produced in excess, and its native bovine (48E2+12E3BP) counterpart. The results of the present study provide a rational basis for resolving apparent differences between previous models, both obtained using rhE2/E3BP core assemblies where no account was taken of relative E2 and E3BP expression levels. Mathematical modelling predicts that an ‘average’ 48E2+12E3BP core arrangement allows maximum flexibility in assembly, while providing the appropriate balance of bound E1 and E3 enzymes for optimal catalytic efficiency and regulatory fine-tuning. We also show that the rhE2/E3BP and bovine E2/E3BP cores bind E3s with a 2:1 stoichiometry, and propose that mammalian PDC comprises a heterogeneous population of assemblies incorporating a network of E3 (and possibly E1) cross-bridges above the core surface.This work was partly supported by EPSRC (under grants GR/R99393/01 and EP/C015452/1)

作物近縁野生種の収集と調査, 1997 : 2. 本州中部地域におけるアズキおよびダイズ近縁野生種

An exploration was conducted from 15^ to 24^ October to find growing sites of wild azuki bean (Vigna angularis var. nipponensis) and wild soybean (Glycine soja) and to collect seeds, nodules and herbarium specimens. A total of 44 samples from 26 sites were collected, which consist of 16 samples of wild soybean, 9 of wild azuki bean, 8 of weedy azuki bean, 4 of cultivated azuki bean, 4 of wild and weedy azuki complex and 3 of Vicia sp. Among 26 sites, wild soybean and wild azuki bean were sympatric in 6 sites. In some cases (Site No. 69, 73, 74, 75, 76, 79, 80, 83), variation in seed color and/or pod color were observed for wild and weedy azuki bean populations. In Tottori site (Site No. 72 and 73), wild, weedy and cultivated azuki bean were growing sympatrically. These sites may be interesting to study gene flow and origin of "weedy" type of azuki. After seed increase, accessions will be preserved in the MAFF genebank. Herbarium specimens are deposited in the MAFF genebank herbarium

Structure of the Helicase Domain of DNA Polymerase Theta Reveals a Possible Role in the Microhomology-Mediated End-Joining Pathway

DNA polymerase theta (Polθ) has been identified as a crucial alternative non-homologous end-joining factor in mammalian cells. Polθ is upregulated in a range of cancer cell types defective in homologous recombination, and knockdown has been shown to inhibit cell survival in a subset of these, making it an attractive target for cancer treatment. We present crystal structures of the helicase domain of human Polθ in the presence and absence of bound nucleotides, and a characterization of its DNA-binding and DNA-stimulated ATPase activities. Comparisons with related helicases from the Hel308 family identify several unique features. Polθ exists as a tetramer both in the crystals and in solution. We propose a model for DNA binding to the Polθ helicase domain in the context of the Polθ tetramer, which suggests a role for the helicase domain in strand annealing of DNA templates for subsequent processing by the polymerase domain

Peraturan Bersama Menteri Agama dan Menteri dalam Negeri Nomor 08 dan 09 Tahun 2006 Tentang Pendirian Rumah Ibadat (Kajian dalam Perspektif Hak Asasi Manusia )

Pundamental 1945 Constitution as the rule has been set explicitly for religious freedom in the run by followers, who belong to one human rights has, but in reality the persecution of religious life and often inevitable. Between religious persecution can come from various directions such as harassing each other, mutual intimidate(violence) and that most often occurs between religious persecution that is prihal establishment synagogue. Therefore, in terms of the establishment of the synagogue there must be government intervention to regulate it, if in this case given the freedom and without any clear rules, the sectarian conflict will not be able to avoid. One step from the government to avoid conflict in the establishment of the synagogue is to be issued the Joint Decree of the Minister of Religious Affairs and the Minister of Home Affairs Number 08 and Number 09 Year 2006 About the Construction of Houses of Worship that aims to create harmony and peace between religious and have certainty Strong law