Different mechanisms of the biochemical adaptation of mycelial fungi to temperature stress: Changes in the cytosol carbohydrate composition

The effect of temperature stress on the cytosol carbohydrate composition of fungi belonging to various systematic groups was investigated. InMucorales representatives (subkingdomEomycota, phylumArchetnycota, classZygomycetes), adaptation to hypo- and hyperthermia occurs via the regulation of trehalose synthesis, although inositol is also involved in these processes inBlakeslea trispora. InAscomycota (subkingdomNeomycota), oversynthesis of mannitol and glycerol occurs under hypothermia, whereas oversynthesis of trehalose and inositol takes place under hyperthermia.Basidiomycota (subkingdomNeomycota) use two pathways of biochemical adaptation, depending on the cytosol carbohydrate composition. In the absence of sucrose, glycerol and arabitol are involved in the adaptation to hyperthermia; trehalose accumulates under hypothermic conditions (type I of regulation). Type II regulation (revealed inPleurotus ostreatus) involves sucrose rather than glycerol or arabitol. The data obtained are discussed in terms of fungal systematics and phylogeny.     

稀释液中添加海藻糖对山羊精子功能和膜完整性的影响

选用5只年龄为3～4岁的波尔山羊公羊研究在稀释液中添加海藻糖对山羊精子功能和膜完整性的影响。山羊精子分别用含6.6 mmol/L、13.2 mmol/L、19.8 mmol/L、26.4 mmol/L、39.6 mmol/L、52.9 mmol/L、66.1mmol/L、79.3 mmol/L的不同海藻糖的Tris-柠檬酸-葡糖糖(TCG)稀释液(卵黄:18%;甘油:6%)稀释和冷冻。结果表明:39.6 mmol/L、52.9 mmol/L、66.1 mmol/L、79.3 mmol/L组降温后的精子活率显著(P<0.05)降低;52.9 mmol/L、66.1 mmol/L、79.3 mmol/L组降温后的精子畸形率和39.6 mmol/L组降温后的膨胀精子率显著(P<0.05)提高。26.4 mmol/L组和39.6 mmol/L组冻融后的精子活率显著(P<0.05)高于对照组;66.1mmol/L和79.3 mmol/L组冻融后的精子活率、畸形率分别显著(P<0.05)低于和高于对照组。19.8 mmol/L、26.4 mmol/L、39.6 mmol/L组冻融后精子获能率显著(P<0.05)低于对照组。39.6 mmol/L组冻融后顶体完整率和膨胀精子率显著(P<0.05)高于对照组,而66.1 mmol/L组和79.3 mmol/L组显著(P<0.05)低于对照组。39.6 mmol/L组的受胎率显著(P<0.05)高于对照组,而66.1mmol/L组和79.3 mmol/L组的受胎率显著(P<0.05)低于对照组。结果表明,在含18%的卵黄(v/v)、6%甘油(v/v)的TCG稀释液中,添加适宜浓度(26.4mmol/L和39.6 mmol/L)海藻糖,可显著提高山羊精子功能和膜的完整性。     

定向进化提高来源于Arthrobacter ramosus的MTHase的热稳定性

麦芽寡糖基海藻糖水解酶(mahosyhrehalose hydrolase,MTHase)是以淀粉或麦芽糊精为底物制备海藻糖的关键酶之一.来源于Arthrobacter ramosus的MTHase,表达量好,比活高,但热稳定性差,限制了其工业化应用.采用定向进化技术,筛选得到L137M和A216T两个突变体,在60℃...     

UV-B照射培养对酵母菌生理活性物质的影响

研究了UV-B照射培养过程中酵母细胞内各种生理活性物质的变化。实验结果显示,UV-B照射培养过程中,酵母细胞中RNA、蛋白质、海藻糖、麦角甾醇和葡聚糖含量均有不同程度的提高,其中RNA含量由0h的8.94%增加到72h的9.88%;蛋白质含量在72h时达到最大值,比培养初期提高0.28%;海藻糖在60h达到最高值,约为113.9mg·g-1酵母;麦角甾醇含量在84h达到最大值为15.43mg·g-1酵母;葡聚糖在72h时的含量占细胞壁干重的22.60%。而酵母细胞中谷胱甘肽的含量和超氧化物歧化酶活性则均呈下降趋势。说明UV-B照射对酵母生长产生较大影响,多种生理活性物质的含量出现不同变化。     

Enzymatic processes for the purification of trehalose

A dual‐enzyme process aiming at facilitating the purification of trehalose from maltose is reported in this study. Enzymatic conversion of maltose to trehalose usually leads to the presence of significant amount of glucose, by‐product of the reaction, and unreacted maltose. To facilitate the separation of trehalose from glucose and unreacted maltose, sequential conversion of maltose to glucose and glucose to gluconic acid under the catalysis of glucoamylase and glucose oxidase, respectively, is studied. This study focuses on the hydrolysis of maltose with immobilized glucoamylase on Eupergit® C and CM Sepharose. CM Sepharose exhibited a higher protein adsorption capacity, 49.35 ± 1.43 mg/g, and was thus selected as carrier for the immobilization of glucoamylase. The optimal reaction temperature and reaction pH of the immobilized glucoamylase for maltose hydrolysis were identified as 40°C and 4.0, respectively. Under such conditions, the unreacted maltose in the product stream of trehalose synthase‐catalyzed reaction was completely converted to glucose within 35 min, without detectable trehalose degradation. The conversion of maltose to glucose could be maintained at 0.92 even after 80 cycles in repeated‐batch operations. It was also demonstrated that glucose thus generated could be readily oxidized into gluconic acid, which can be easily separated from trehalose. We thus believe the proposed process of maltose hydrolysis with immobilized glucoamylase, in conjunction with trehalose synthase‐catalyzed isomerization and glucose oxidase‐catalyzed oxidation, is promising for the production and purification of trehalose on industrial scales. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2013     

Induction of the differentiation of human HL-60 promyelocytic leukemia cell line by succinoyl trehalose lipids

Four analogs of succinoyl trehalose lipid-3 (STL-3)with saturated even-number or odd-number carbonchains, and unsaturated or halogenated fatty acidswere examined for their ability to inhibit the growthand induce the differentiation of HL-60 humanpromyelocytic leukemia cells. The optimalconcentration of STL-3 at which such activities wererecognized was closed to the critical micelleconcentration of STL-3. Analog of STL-3 witheven-number or odd-number carbon chain and unsaturatedfatty acids strongly inhibited growth and induced thedifferentiation of HL-60 cells, as evaluated in termsof nitroblue tetrazilium-reducing activity and theappearance of the CD36 antigen. An analog of STL-3with halogenated fatty acids significantly inhibitedproliferation but only induced the differentiation ofHL-60 cells. Our results indicate that the effects ofSTL-3 and its analogs on HL-60 cells depend on thestructure of the hydrophobic moiety of STL-3.These authors contributed equally to this work     

硫矿硫化叶菌MTSase和MTHase基因的克隆与表达

从嗜热硫矿硫化叶菌(Sulfolobussolfataricus)ATCC35092的基因组中用PCR方法扩增得到编码MTSase和MTSase的基因,分别将其插入原核表达载体pTrc99a中,并转入大肠杆菌BL21(DE3),进行诱导表达。MTSase和MTHase酶活产率达到了31.3U/g(wetcell)和403U/g(wetcell)。在75℃,pH5.0条件下,两酶联合作用转化淀粉生产海藻糖,当淀粉浓度为15%,DE值为10时,海藻糖转化率最高为53.6%。     

栗精碱对棉铃虫中肠组织可溶型海藻糖酶的抑制效应

【目的】研究一种植物源化合物栗精碱对棉铃虫Helicoverpa armigera(Hübner)5龄幼虫中肠组织可溶型海藻糖酶的活体和离体抑制效果,初步探索其抑制作用。【方法】经过离子交换层析和疏水层析从棉铃虫5龄幼虫中肠组织分离纯化得到可溶型海藻糖酶,通过凝胶过滤层析确定其分子量。分别用不同浓度的抑制剂进行离体和活体抑制研究,得出栗精碱对可溶型海藻糖酶的抑制率和抑制中浓度(IC50),并采用Lineweaver-Burk和Dixon作图法分析抑制类型。【结果】棉铃虫中肠可溶型海藻糖酶比活力为2.022 U/mg,回收率为67.57%,纯化倍数为23.84,分子量约为67 k Da。离体抑制研究表明,3.75,7.5,15和30μmol/L的栗精碱对可溶型海藻糖酶的抑制率分别为42.00%,50.30%,58.32%和67.33%;抑制中浓度(IC50)为7.32μmol/L。通过LineweaverBurk和Dixon作图法,确定栗精碱是棉铃虫中肠组织可溶型海藻糖酶的一种有效的竞争性抑制剂,Ki值为4.9μmol/L。活体抑制研究表明,分别注射浓度为30,15和7.5μmol/L栗精碱10和20 h后,棉铃虫中肠可溶型海藻糖酶活性被显著抑制(P0.05)。相应地,注射后随着时间的延长,血淋巴海藻糖含量连续增加。而注射栗精碱20 h后,血淋巴葡萄糖浓度显著下降,使棉铃虫的能量供给出现障碍。【结论】结果表明,栗精碱是棉铃虫中肠可溶型海藻糖酶的一种有效的抑制剂,可为下一步开发有效的棉铃虫杀虫剂提供理论支持。     

Trehalose and trehalose-hydrolyzing enzyme in the haemolymph of Locusta migratoria infected with Metarhizium anisopliae strain CQMa102

Topical application of the Metarhizium anisopliae var. acridum specialist strain CQMa 102 to the locust Locusta migratoria manilensis results in changes of the concentrations of trehalose and glucose in the haemolymph. Micrographs of the locust haemolymph shows Metarhizium anisopliae can effectivly penetrate the external skeleton of locust and after 2 days infection, the hyphae body will appear in the haemolymph of infected insects. The time in decrease of trehalose concentration coincided with that in increase of trehalose-hydrolysing enzyme activity in the haemolymph of the fungus-infected insects. Overlay gel analysis indicated there was considerably more trehalose-hydrolysing activity in the haemolymph of locusts infected by fungus than in controls. A comparable isoform was identified in in vitro culture of the fungus, suggesting a fungal origin for the in vivo enzyme. Haemolymph trehalose decreased significantly during mycosis of locusts by M. anisopliae. All these results suggested that this fungus may take advantage of competing nutrient utilization against the insect by its trehalose-hydrolyzing enzyme secretion. It may provide fundamental knowledge for fungal pathogenesis.     

Feedforward Regulation of Glucose Metabolism by Steroid Hormones Drives a Developmental Transition in Drosophila

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