Anti-oxidative effects of silkworm storage protein 1 in HeLa cell

Silkworm hemolymph contains unique proteins that exhibit anti-apoptotic activity in mammalian cells. Among them, 30 K protein, which is one of the major anti-apoptotic molecules in silkworm hemolymph, has been well investigated. However, little is known about the biological functions of storage protein 1 (SP1), another main protein in silkworm hemolymph. In this study, the anti-apoptotic and anti-oxidative activities of SP1 were analyzed. A stable cell line expressing SP1 was constructed, which showed strong anti-apoptotic effect induced by staurosporine treatment. In addition, the cell line exhibited resistance to oxidative stress caused by hydrogen peroxide. For practical applications of SP1, recombinant SP1 was produced in Escherichia coli, and the supplementation of recombinant SP1 into culture medium exhibited anti-apoptotic and anti-oxidative activities. In addition, SP1 was found to be a cell-penetrating protein and localized in the cytosol as well as on the plasma membrane. The findings showed that SP1 itself is not an anti-oxidant; rather, it mediates intracellular anti-oxidative activity. In conclusion, the cellular resistance of SP1 to apoptosis and oxidative stress will provide a new strategy that could be utilized in the bio-industry for the production of biologics as well as for the development of anti-aging cosmetics.     

Mobilisation of the major stored reserves in the embryo of fenugreek (Trigonella foenum-graecum L., Leguminosae), and correlated enzyme activities

Changes in total nitrogen, soluble amino nitrogen, lipid and phytate contents, and in the activities of proteinase (pH 7.0), isocitrate lyase and phytase were followed in the endosperm, cotyledons, and axis during germination of fenugreek seeds and subsequent growth of the seedlings. The endosperm is comprised largely of cell-wall galactomannans: the majority of the seed total nitrogen, lipid and phytate (5%, 8%, 0.44% of seed dry weight respectively) is localised within the cotyledons as stored reserves. Germination is completed after 10–14 h from the start of imbibition, but the major reserves are not mobilised during the first 24 h. Then the total nitrogen content of the cotyledons starts to decrease and that of the axis increases; there is a concomitant accumulation of soluble amino nitrogen in both cotyledons and axis. An increase in proteinase activity in the cotyledons correlates well with the depletion of total nitrogen therein. Depletion of lipid and phytate reserves in the different seed tissues constitutes a late event, occurring after 50 h from the start of imbibition, and is coincident with the final disintegration of the endosperm tissue. The depletion of phytate and stored lipids is accompanied by an increase in phytase and isocitrate lyase activity. It appears that the products of lipid hydrolysis are converted by gluconeogenesis to serve as the major source of sugars for the growing axis after the endosperm galactomannan has been completely mobilised.     

Stability of the intra- and extracellular toxins of Prymnesium parvum using a microalgal bioassay

Prymnesium parvum produces a variety of toxic compounds, which affect other algae, grazers and organisms at higher trophic levels. Here we provide the method for development of a sensitive algal bioassay using a microalgal target, Teleaulax acuta, to measure strain variability in P. parvum toxicity, as well as the temporal stability of both the intracellular and the extracellular lytic compounds of P. parvum. We show high strain variation in toxicities after 3 h incubation with LC₅₀s ranging from 24 to 223 × 10³ cells ml⁻¹. Most importantly we prove the necessity of testing physico-chemical properties of P. parvum toxins before attempting to isolate and characterize them. The extracellular toxin in the supernatant is highly unstable, and it loses significant lytic effects after 3 days despite storage at −20 °C and after only 24 h stored at 4 °C. However, when stored at −80 °C, lytic activity is more easily maintained. Reducing oxidation by storing the supernatant with no headspace in the vials significantly slowed loss of activity when stored at 4 °C. We show that the lytic activity of the intracellular toxins, when released by sonication, is not as high as the extracellular toxins, however the stability of the intracellular toxins when kept as a cell pellet at −20 °C is excellent, which proves this is a sufficient storage method for less than 3 months. Our results provide an ecologically appropriate algal bioassay to quantify lytic activity of P. parvum toxins and we have advanced our knowledge of how to handle and store the toxins from P. parvum so as to maintain biologically relevant toxicity.     

Arylsulfatase K,a Novel Lysosomal Sulfatase

The human sulfatase family has 17 members, 13 of which have been characterized biochemically. These enzymes specifically hydrolyze sulfate esters in glycosaminoglycans, sulfolipids, or steroid sulfates, thereby playing key roles in cellular degradation, cell signaling, and hormone regulation. The loss of sulfatase activity has been linked to severe pathophysiological conditions such as lysosomal storage disorders, developmental abnormalities, or cancer. A novel member of this family, arylsulfatase K (ARSK), was identified bioinformatically through its conserved sulfatase signature sequence directing posttranslational generation of the catalytic formylglycine residue in sulfatases. However, overall sequence identity of ARSK with other human sulfatases is low (18–22%). Here we demonstrate that ARSK indeed shows desulfation activity toward arylsulfate pseudosubstrates. When expressed in human cells, ARSK was detected as a 68-kDa glycoprotein carrying at least four N-glycans of both the complex and high-mannose type. Purified ARSK turned over p-nitrocatechol and p-nitrophenyl sulfate. This activity was dependent on cysteine 80, which was verified to undergo conversion to formylglycine. Kinetic parameters were similar to those of several lysosomal sulfatases involved in degradation of sulfated glycosaminoglycans. An acidic pH optimum (∼4.6) and colocalization with LAMP1 verified lysosomal functioning of ARSK. Further, it carries mannose 6-phosphate, indicating lysosomal sorting via mannose 6-phosphate receptors. ARSK mRNA expression was found in all tissues tested, suggesting a ubiquitous physiological substrate and a so far non-classified lysosomal storage disorder in the case of ARSK deficiency, as shown before for all other lysosomal sulfatases.     

白木香种子脱水耐性的发育变化及贮藏特性

研究白木香种子发育进程中种子性状、萌发能力和脱水耐性的变化,以及不同光温条件对种子萌发的影响和种子的贮藏特性。结果表明：白木香种子在花后78d获得最大干重,进入生理成熟期,此时萌发率接近最大值;胚在花后57～85d,脱水耐性逐渐增强,并在花后85d获得最大脱水耐性。种子萌发的适宜温度范围为25℃-35℃,光照对种子萌发有一定的抑制作用。新鲜自木香种子（含水量27．45％）在4℃低温条件下贮藏1个月后萌发率仅为30％左右,而含水量7．38％的干燥种子在4℃低温条件下贮藏120d,萌发率仍有53．33％,因此,4℃低温和适度脱水有利于种子短期贮藏。白木香种子能忍耐一定程度的脱水,但干燥至含水量7．50％以下时种子会发生损伤,因此推测白木香种子是一种中间性种子。     

Competition and overlap of Oryzaephilus surinamensis and Plodia interpunctella populations under condition of stored date fruits

Plodia interpunctella and Oryzaephilus surinamensis are found in food storehouses including dates and palm storages. The current study aimed to determine competition and overlap potentials of the two pests of date fruits. Time series models were used to study two species populations and logistic growth model to estimate the effect of density of the species. The results revealed the environmental capacities of O. surinamensis and P. interpunctella were 433 and 1610 (maximum number per 20 g), respectively, and the population growth rates (r) were 1.2 and 1.3, respectively. Ecological balances of the two species were close to each other from the first to the third week. The population of O. surinamensis decreased in the fourth week of the competition. The highest population balance of the two species was in the 14th week. The potential of exploitable ecological niches (e_ij) and the amount of non-exploited ecological niches by any species (z_ij) for O. surinamensis was higher than for P. interpunctella from the 8th week untill the end of sampling period. The overlap of ecological niches in the two species (D) ranged from 0.94 to 1, indicating a complete overlap of temporal activity in the two populations on date palm. The current results of this study can be used by integrated pest management specialists. Information over the effects of species competition on population dynamics and their coexistence can be used to predict population status and to adopt simple pest control methods.     

Characterization of glycolytic initial metabolites and enzyme activities in developing sunflower (Helianthus annuus L.) seeds

Unlike other oilseeds (e.g. Arabidopsis), developing sunflower seeds do not accumulate a lot of starch and they rely on the sucrose that comes from the mother plant to synthesise lipid precursors. Between 10 and 25 days after flowering (DAF), when sunflower seeds form and complete the main period of storage lipid synthesis, the sucrose content of seeds is relatively constant. By contrast, the glucose and fructose content falls from day 20 after flowering and it is always lower than that of sucrose, with glucose being the minor sugar at the end of the seed formation. By studying the apparent kinetic parameters and the activity of glycolytic enzymes in vitro, it is evident that all the components of the glycolytic pathway are present in the crude seed extract. However, in isolated plastids important enzymatic activities are missing, such as the glyceraldehyde-3-phosphate dehydrogenase, involved in the conversion of glyceraldehyde 3-phosphate into 1,3-biphospho-glycerate, or the enolase that converts 2-phosphoglycerate into phosphoenolpyruvate. Hence, phosphoenolpyruvate or one of its derivatives, like pyruvate and malate from the cytosol, may be the primary carbon sources for lipid biosynthesis. Accordingly, the glucose-6-P imported into the plastid is likely to be used in the pentose phosphate pathway to produce the reducing power for lipid biosynthesis in the form of NADPH. Data from crude seed extracts indicate that enolase activity increased during seed formation, from 16 days after flowering, and that this activity was well correlated with the period of storage lipid synthesis. In addition, while the presence of some glycolytic enzymes increased during lipid synthesis, others decreased, remained constant, or displayed irregular temporal behaviour.     

新疆冬春麦区小麦地方品种贮藏蛋白遗传多样性研究

采用SDS-聚丙烯酰胺凝胶电泳（SDS-PAGE）对236份新疆小麦地方品种的高分子量麦谷蛋白亚基（HMW-GS）的组成进行了分析。结果表明：Glu-Ⅰ位点共有19种等位基因,其中Glu-Al位点3种,Glu-Bl位点7种,Glu—D1住点9种;亚基null、7＋8、2＋12在各自的位点上出现频率最高,分别达到91．95％、85．17％、80．93％;亚基组成类型共有21种,主要为null／7＋8／2＋12,频率达70．34％;同时筛选出33份含有1、2^＊、13＋16、14＋15、5＋10、1．5＋10、174-18等优质亚基的材料,可作为优质基因源。利用酸性聚丙烯酰胺凝胶电泳（A-PAGE）对其中的65份地方品种进行醇溶蛋白多样性分析。结果表明：电泳出现64条迁移率不同的谱带,构成65种组合,其中ω区出现的谱带最多,达17条;其次是β和γ区各16条,α区出现的谱带数最少,为15条。从每条谱带在65份材料中出现的频率看,总的变异范围为1．54％～93．85％;α、β、γ和ω4个分区多样性指数（H1）分别为0．498、0．386、0．523和0．348,表明新疆麦区小麦地方品种贮藏蛋白位点存在丰富的遗传多样性。     

Rheology and gelation of water-insoluble dextran from Leuconostoc mesenteroides NRRL B-523

Dynamic oscillatory testing has been used to study the rheology of water-insoluble dextran. The rheological properties (storage and loss moduli) of dextran gel were measured and dextran was found to be neither a strong gel nor a weak gel, but an entanglement network at a concentration of 250 mg/ml. The extent of gelation, illustrated by the gel elastic modulus G′, is found to decrease with increasing concentration of calcium ions. This was confirmed by shift of crossover frequencies towards higher values on the dynamic spectra and lower yield stress τ values obtained from stress ramp experiments. Finally, a comparison between gelation of dextran and alginate (a similar biopolymer) was made for clear understanding of effect of calcium ions on the dextran gelation.