Crystal structures and structural stabilities of the disulfide bond-deficient soybean proglycinin mutants C12G and C88S

The constituent subunits of seed storage protein 11S globulin have two disulfide bonds that are common among 11S globulins from legume and nonlegume seeds. In the case of the A1aB1b subunit of soybean 11S globulin, glycinin, Cys12-Cys45 and Cys88-Cys298 are observed by X-ray crystallography. The significance of these two disulfide bonds for structural stability was investigated by mutagenesis of Cys12 to Gly and of Cys88 to Ser. The disulfide bond-deficient mutants C12G and C88S could form the correct conformations identical to that of the wild-type proglycinin except in the vicinities of the mutation sites C12 and C88 as shown by their crystal structures. Thermal stability monitored by differential scanning calorimetry of the mutants indicated that the contribution of these disulfide bonds to the thermal stability of proglycinin A1aB1b is low, although there is a small difference in the extent of the contribution between the two disulfide bonds (Cys12-Cys45 > Cys88-Cys298). The contribution of Cys88-Cys298 to the resistance of proglycinin A1aB1b to proteinase digestion is higher than that of Cys12-Cys45. Possible effects of structure on the different properties of C12G and C88S are discussed.     

Formation and chemical structures of acid-soluble lignin II: reaction of aromatic nuclei model compounds with xylan in the presence of a counterpart for condensation,and behavior of lignin model compounds with guaiacyl and syringyl nuclei in 72% sulfuric acid

To elucidate the formation and chemical structures of water-soluble material in acid-soluble lignin (ASL), lignin aromatic nuclei model compounds of creosol (I) and 5-methoxycreosol (II) were reacted with xylose or xylan in the presence of apocynol as a counterpart for condensation in 72% sulfuric acid (SA). The reaction of I gave mainly condensation product. However, the condensation reaction of II with apocynol was suppressed because of steric hindrance from the methoxyl group, and II yielded a C-xyloside after refluxing in 3% SA together with condensation products. To obtain information on CHCl₃-soluble material in ASL, model compounds of arylglycerol--aryl ethers with guaiacyl (VIII) and syringyl (X) nuclei were treated by the Klason procedure. VIII gave only insoluble polymerized product, while X gave insoluble polymerized product and CHCl₃-soluble low molecular weight products, which were dissolved in 3% SA. These results prove earlier views that water-soluble material in ASL consists of condensation products formed from syringyl lignin and monosaccharide units in hemicellulose. In addition, the CHCl₃-soluble material in ASL appears to be composed of low molecular weight degradation products from SA treatment of Klason lignin with the syringyl nucleus.Part of this report was presented at the 51st Annual Meeting of the Japan Wood Research Society, Tokyo, April 2001 and at the 47th Lignin Symposium, Fukuoka, October 2002, and was reviewed in Mokuzai Gakkaishi (2002) 48:55–62     

High level expression and purification of bioactive bovine interleukin-18 using a baculovirus system

Bioactive recombinant bovine interleukin-18 (rboIL-18) was expressed using a baculovirus system. Normally, IL-18 is translated as a precursor form of a 24kDa polypeptide and processed by IL-1beta converting enzyme (ICE) to a mature bioactive form of 18kDa protein. Hence, to express active form IL-18, we constructed two recombinant baculoviruses containing boIL-18 and human ICE (hICE) genes, respectively, and superinfected these viruses into insect cells. Superinfection of both recombinant viruses into the cells resulted in the expression of a 24kDa precursor form and an 18kDa mature form detectable in the supernatant by immunoblotting using anti-porcine IL-18 antibody. Culture supernatant from the superinfected cells showed a synergistic effect with recombinant boIL-12 for production of interferon-gamma (IFN-gamma) in bovine peripheral mononuclear cells. By addition of histidine hexamer at the C-terminal of boIL-18, the mature IL-18 was purified. Bioactivity remained after purification.     

Intraspecific Evolution of <Emphasis Type="Italic">Rhizoctonia solani</Emphasis> AG-1 IA Associated with Soybean and Rice in Brazil based on Polymorphisms at the ITS-5.8S rDNA Operon

Rhizoctonia solani AG-1 IA causes leaf blight on soybean and rice. Despite the fact that R. solani AG-1 IA is a major pathogen affecting soybean and rice in Brazil and elsewhere in the world, little information is available on its genetic diversity and evolution. This study was an attempt to reveal the origin, and the patterns of movement and amplification of epidemiologically significant genotypes of R. solani AG-1 IA from soybean and rice in Brazil. For inferring intraspecific evolution of R. solani AG-1 IA sampled from soybean and rice, networks of ITS-5.8S rDNA sequencing haplotypes were built using the statistical parsimony algorithm from Clement et al. (2000) Molecular Ecology 9: 1657–1660. Higher haplotype diversity (Nei M 1987, Molecular Evolutionary Genetics Columbia University Press, New york: 512p.) was observed for the Brazilian soybean sample of R. solani AG-1 IA (0.827) in comparison with the rest of the world sample (0.431). Within the south-central American clade (3-2), four haplotypes of R. solani AG-1 IA from Mato Grosso, one from Tocantins, one from Maranhão, and one from Cuba occupied the tips of the network, indicating recent origin. The putative ancestral haplotypes had probably originated either from Mato Grosso or Maranhão States. While 16 distinct haplotypes were found in a sample of 32 soybean isolates of the pathogen, the entire rice sample (n=20) was represented by a single haplotype (haplotype 5), with a worldwide distribution. The results from nested-cladistic analysis indicated restricted gene flow with isolation by distance (or restricted dispersal by distance in nonsexual species) for the south-central American clade (3-2), mainly composed by soybean haplotypes.     

Aerial Spraying of Neemazal®-T/S Against the Forest Cockchafer (Melolontha hippocastani,Col.: Scarabaeidae) in South-West Germany: The Effects of Two Field Trials Performed in 2007 and 2008 on Local Populations

Journal of Plant Diseases and Protection - The forest cockchafer Melolontha hippocastani is a serious forest pest whose infestation levels have been increasing in South-West Germany since the...     

Growth characteristics,phytate contents,and coagulation properties of soymilk from a low-phytate Japanese soybean (Glycine max (L.) Merr.) line

The phytic acid (myo-inositol hexakisphosphate or InsP₆) content of seed crops is important to their nutritional quality. Since it represents 75?±?10% of the total seed phosphorus (P), phytic acid is also important regarding the management of P in agricultural production. A low-phytate F5 line, No. T-2-250-4-20, was selected from the progeny of a cross between the low-phytate soybean line CX1834 and the Japanese commercial cultivar Tanbakuro. This line and its parents were grown in a field nursery, and the growth characteristics, phytate accumulation, and processing suitability for tofu were evaluated. At full maturity, the weight of seeds per plant of line T-2-250-4-20 was 5.2- and 1.3-fold higher than that of CX1834 and Tanbakuro, respectively. The amount of phytate-phosphorus as a percentage of the total P content in seeds was 23% in line T-2-250-4-20-34, 30% in CX1834, and 69% in Tanbakuro. No significant difference was observed among the three cultivars/lines in their seed magnesium (Mg), potassium (K), crude protein, and sugar. However, the calcium (Ca), crude fat and ash contents in seeds of line T-2-250-4-20-34 and Tanbakuro was lowered compared to that of CX1834. The breaking stress of tofu was estimated employing a rheometer with a decreasing concentration of the coagulant magnesium chloride (MgCl₂), starting at 15.7?mmol?L^?1. In tofu made from Tanbakuro, the concentration of MgCl₂ required to achieve the maximum breaking stress was 12.6?mmol?L^?1; however, it was 9.5?mmol?L^?1 for tofu made from T-2-250-4-20-34 and CX1834. The tofu made from Tanbakuro was soft and broke at 6.3?mmol?L^?1 MgCl₂, but, in line T-2-250-4-20-34, harder tofu was made with lower MgCl₂ concentrations. No difference was observed among the cultivars/lines in the SDS-PAGE patterns of protein in soymilk. These results indicate that we have developed a low-phytate soybean with adequate productivity, and confirmed that tofu made from the low-phytate T-2-250-4-20-34 soybean becomes coagulated and harder at a lower MgCl₂ concentration than that from high-phytate soybean cultivars.     

Insulin-resistant diabetes due to a point mutation that prevents insulin proreceptor processing

A point mutation in the human insulin receptor gene in a patient with type A insulin resistance alters the amino acid sequence within the tetrabasic processing site of the proreceptor molecule from Arg-Lys-Arg-Arg to Arg-Lys-Arg-Ser. Epstein-Barr virus-transformed lymphocytes from this patient synthesize an insulin receptor precursor that is normally glycosylated and inserted into the plasma membrane but is not cleaved to mature alpha and beta subunits. Insulin binding to these cells is severely reduced but can be increased about fivefold by gentle treatment with trypsin, accompanied by the appearance of normal alpha subunits. These results indicate that proteolysis of the proreceptor is necessary for its normal full insulin-binding sensitivity and signal-transducing activity and that a cellular protease that is more stringent in its specificity than trypsin is required to process the receptor precursor.     

An assessment of genetic diversity in wild and captive populations of endangered Japanese bitterling <Emphasis Type="Italic">Tanakia tanago</Emphasis> (Cyprinidae) using amplified fragment length polymorphism (AFLP) markers

ABSTRACT:   The Japanese bitterling Tanakia tanago (Cyprinidae) is on the verge of extinction in the wild, placing great importance on captive breeding programs for current conservation of the species. However, the loss of genetic diversity during captive breeding is an ongoing matter of concern. Since some captive populations have been almost monomorphic in mitochondrial DNA (mtDNA), this hampers assessments of their genetic diversity during captive breeding. To more accurately assess their genetic diversity, one wild and three captive populations were examined using amplified fragment length polymorphism (AFLP) markers. Estimates of average heterozygosity and nucleotide diversity ranged 0.0479–0.1920 and 0.0023–0.0088, respectively, enabling comparison of genetic diversity among the wild and captive populations, and among year-classes of captive populations. Significant differences in numbers of amplified fragments and proportions of polymorphic fragments were observed among year-classes of all populations. The indices of genetic diversity calculated from AFLP seemed to be, however, less sensitive to weak bottlenecks. No continuous decrease in genetic diversity in nuclear DNA was detected in presently captive populations. This supports the possibility of re-introduction of the captive populations into the original habitats, although survival and reproductive ability in the wild must be taken into consideration.     

Permeation of carboxylates as ionic compounds into fish meats by soaking and its osmotic dehydrating effect

Permeability of Na-gluconate, Na-malate and Na-acetate into fish-meat strips by soaking, and their osmotic dehydrating effects were investigated in comparison with those of sodium chloride (NaCl) and sorbitol. Carboxylic acids, anion species of carboxylates, were less permeable into soaked meats than chlorine of NaCl, and their permeability was similar to that of sorbitol. However, the permeation of sodium, cation species of the carboxylates and NaCl, was promoted with the concentration of free sodium ion dissociated in soaking solutions. The ratio of sodium to the anion species in soaked meats was varied, depending on the numbers of the sodium atom in the molecules as well as the dissociation degree of the ionic compounds in the soaking solution. Furthermore, the osmotic dehydrating effects of Na-gluconate and Na-malate were analogous with that of sorbitol, and higher than those of NaCl and Na-acetate. These results indicated that the permeation characteristics and the osmotic dehydrating effects of the carboxylates were associated with their ionization in soaking solution as well as their molecular weights.