Cysteine [2,4] Disulfide Bond as a New Modifiable Site of α-Conotoxin TxIB

α-Conotoxin TxIB, a selective antagonist of α6/α3β2β3 nicotinic acetylcholine receptor, could be a potential therapeutic agent for addiction and Parkinson’s disease. As a peptide with a complex pharmacophoric conformation, it is important and difficult to find a modifiable site which can be modified effectively and efficiently without activity loss. In this study, three xylene scaffolds were individually reacted with one pair of the cysteine residues ([1,3] or [2,4]), and iodine oxidation was used to form a disulfide bond between the other pair. Overall, six analogs were synthesized with moderate isolated yields from 55% to 65%, which is four times higher than the traditional two-step oxidation with orthogonal protection on cysteines. The cysteine [2,4] modified analogs, with higher stability in human serum than native TxIB, showed obvious inhibitory effect and selectivity on α6/α3β2β3 nicotinic acetylcholine receptors (nAChRs), which was 100 times more than the cysteine [1,3] modified ones. This result demonstrated that the cysteine [2,4] disulfide bond is a new modifiable site of TxIB, and further modification can be a simple and feasible strategy for the exploitation and utilization of α-Conotoxin TxIB in drug discovery.     

Distribution of protein disulfide isomerase during maturation of pig oocytes

Oocyte maturation in mammals is characterized by a dramatic reorganization of the endoplasmic reticulum (ER). In mice, the ER forms accumulations in the germinal vesicle (GV) stage and distinctive cortical clusters in metaphase II (MII) of the oocyte. Multiple evidence suggests that this ER distribution is important in preparing the oocyte for Ca²⁺ oscillations, which trigger oocyte activation at fertilization. In this study, we investigated the time course and illustrated the possible functional role of ER distribution during maturation of porcine oocytes by immunostaining with protein disulfide isomerase (PDI). PDI forms clusters in the cytoplasm of oocytes. After immunostaining, PDI clusters were identified throughout the cytoplasm from the GV to metaphase I (MI) stage; however, at the MII stage, the PDI formed large clusters (1–2 µm) in the animal pole around the first polar body. PDI distribution was prevented by bacitracin, a PDI inhibitor. Our experiments indicated that, during porcine oocyte maturation, PDI undergoes a dramatic reorganization. This characteristic distribution is different from that in the mouse oocyte. Moreover, our study suggested that formation of PDI clusters in the animal pole is a specific characteristic of matured porcine oocytes.     

吡咯伯克霍尔德菌WY6-5产二甲基二硫对储藏期花生黄曲霉及毒素的抑制作用

【目的】验证吡咯伯克霍尔德菌（Burkholderia pyrrocinia）WY6-5的抑菌作用,评价其对储藏期花生黄曲霉（Aspergillus flavus）及毒素的防治效果,分析抑菌作用机制,鉴定活性物质,并检测其最低抑菌浓度,为储藏期真菌病害及毒素的防控提供新材料。【方法】采用非接触培养皿对扣培养法,检测菌株WY6-5对黄曲霉的抑制效果,添加活性炭,验证挥发性物质的抑菌作用;密闭储藏环境,检测WY6-5产二甲基二硫对花生黄曲霉及毒素的抑制效果;收集处理后的花生籽粒,锇酸固定,并进行扫描电镜观察,检测黄曲霉细胞显微结构变化,通过透射电镜观察,检测黄曲霉细胞内部结构的显微差异;购买活性物质标准品,梯度稀释,与黄曲霉菌丝和孢子对扣培养,分析最低抑菌浓度。【结果】吡咯伯克霍尔德菌WY6-5分离自茶园根际土壤,可产生挥发性物质二甲基二硫,并高效抑制黄曲霉的生长,抑菌率达95%以上;同时,在高水活度（a_w）条件下,WY6-5还可抑制储藏期花生黄曲霉和毒素污染;两种水活度下,对照组中发病率高达100%,黄曲霉毒素总含量分别为399.32 μg?kg ^-1（a_w 0.859）和3 143.19 μg?kg ^-1（a_w 0.923）;WY6-5添加组,花生黄曲霉发病率降为2%（a_w 0.859）与21%（a_w 0.923）,毒素含量降为4.86 μg?kg ^-1（a_w 0.859）和121.37 μg?kg ^-1（a_w 0.923）,与对照组相比,对毒素的抑制率达98.78%和96.14%。扫描电镜观察显示,WY6-5产生的挥发性气体能抑制黄曲霉孢子的萌发,透射电镜显示,黄曲霉细胞结构未呈现明显损伤。挥发性物质二甲基二硫抑菌作用明显,对黄曲霉菌丝生长的最低抑菌浓度为100 μL?L ^-1(物质体积/培养体积),对孢子萌发的最低抑菌浓度为50 μL?L ^-1(物质体积/培养体积)。【结论】吡咯伯克霍尔德菌WY6-5可产生高效抑菌挥发物二甲基二硫,在低浓度下即可完全抑制黄曲霉菌丝生长和孢子萌发,并能抑制储藏期花生黄曲霉的侵染和黄曲霉毒素的产生,为储藏期真菌病害及毒素的防控提供了新型生物材料。     

热加工过程中鲍鱼腹足蛋白间作用力及其质构特性

为探究蛋白质形成凝胶过程中其化学作用力的变化规律以及与凝胶特性之间的关系,该文以鲍鱼腹足为原料,采用溶液分级提取方法,并结合扫描电镜和红外光谱法,考察热加工中鲍鱼腹足蛋白间作用力及其质构特性的变化情况。结果表明,随着加热温度升高(60、80、100℃),扫描电镜结果显示,鲍鱼腹足中间部位与边缘和过渡部位形成孔洞较小、排列紧密的网络结构,同时红外结果表明,随温度升高,蛋白二级结构发生明显变化,N-H弯曲和C-N伸缩振动较为明显,α-螺旋变为无规则卷曲结构,肌球蛋白疏水性增加,-S-S-形成。此时,对应离子键、氢键含量均呈下降趋势,疏水键相对含量呈先上升后下降趋势,二硫键、非二硫共价键含量呈上升趋势。进一步研究表明,各化学作用力与蛋白凝胶质构特性具有高度相关性。在较低温度下(60℃)离子键、氢键和疏水键对凝胶稳定性起主要作用,此时形成的凝胶较柔软;在较高温度下(80、100℃)二硫键、非二硫共价键为维持凝胶稳定的主要作用力,此时凝胶特性较佳,富有弹性、较好的凝聚性和回复性。该研究为热加工过程中鲍鱼腹足蛋白质变化的机理提供参考依据。     

二甲基甲酰胺对家蚕毒性试验

二甲基甲酰胺对家蚕的毒性较低，只有在很高的浓度食下时才表现出急性中毒死亡，并且随着龄期的增加，家蚕的耐毒性增加，3龄蚕比2龄蚕有更强的耐毒性。低浓度的二甲基甲酰胺（DMF）对家蚕生命率和经济性状都没有显著影响。DMF对家蚕的毒性有积累作用，3龄期连续添毒稀释10倍的DMF原液24h没有观察到急性中毒死亡，但连续添毒90h后，可最终导致家蚕中毒死亡。     

二硫化碳火焰光度气相色谱法的测定研究

使用100 mL全玻璃注射器采集CS2气体,注入HC-6型微量硫气相色谱分析仪,经GDX-104色谱柱分离,应用火焰光度检测器检测。测量的线性检测范围为0.0005~0.855 mg/m3,线性方程为Y=32.67869+1.69828X(r=0.99968),相对标准偏差为2.53%~4.98%,最低检出限为0.0005 mg/m3。     

小麦品种“陕253”PDI基因的克隆、原核表达及蛋白纯化

为进一步研究控制小麦蛋白二硫键形成的关键蛋白即蛋白质二硫键异构酶（PDI）,运用RT-PCR方法克隆出小麦品种＂陕253＂PDI基因的cDNA序列,基因登陆号为HQ911363。序列分析表明,HQ911363全长为1 539 bp,编码512个氨基酸,含有PDI典型的异构酶活性的催化位点-CGHC-和内质网驻留信号肽-KDEL-。构建该基因的原核表达载体,在宿主菌E.coliBL21（DE3）中经IPTG诱导表达融合蛋白,并对表达蛋白进行了纯化。SDS-PAGE及Western-blot检测证实融合蛋白诱导表达并纯化成功。     

二甲基—1,6—辛二烯的环化水合反应研究

３，７－二甲基－１，６－辛二烯（１）在固体酸（分子筛，离子膜和离子交换树脂）作用下与甲酸发生环化水合反应，生成１－（３，３－二甲基环己基）乙醇甲酸酯（４ａ）和３，７，７－三甲基环庚醇甲酸酯（５ａ）的混合物，经皂化反应得到相应的醇（４ｂ和５ｂ）。５，７－二甲基－１，６－辛二烯（２）在同样条件下发生类似的反应，生成３，３，５－三甲基环庚醇甲酸酯（６ａ）经皂化生成３，３，５－三甲基环庚醇（６ｂ）。     

Dimethyl disulfide is an induced systemic resistance elicitor produced by Bacillus cereus C1L

BACKGROUND: Bacillus cereus C1L is a plant growth‐promoting rhizobacterium and can elicit induced systemic resistance (ISR) in plants against necrotrophic pathogens. However, little is known about ISR elicitors produced by B. cereus C1L, and no ISR elicitor has been identified and characterised. Therefore, the objective of this study is to identify volatile ISR elicitor(s) produced by B. cereus C1L. RESULTS: The volatile metabolites produced by B. cereus C1L were extracted, separated and identified by solid‐phase microextraction, gas chromatography and mass spectrometry. Dimethyl disulfide (DMDS) was the only separated metabolite being determined. Afterwards, application of DMDS by means of soil drench significantly protected tobacco and corn plants against Botrytis cinerea and Cochliobolus heterostrophus, respectively, under greenhouse conditions. The results reveal that DMDS could play an important role in ISR by B. cereus C1L. CONCLUSION: This is the first report of DMDS as an elicitor produced by an ISR‐eliciting B. cereus strain and its ability to suppress plant fungal diseases under greenhouse conditions. It is suggested that DMDS has potential for practical use in controlling plant foliar diseases besides soil fumigation. Copyright © 2012 Society of Chemical Industry     

橡胶初加工废水污染土壤的微生物固定化修复

橡胶初加工废水产生大量富含高有机质,并具有高粘度、酸性和高有机物等特点。因其处理成本较高且经常存在厌氧处理失效导致后端废水处理不达标、甚至有较多废水直接排出而污染了周边土壤,但目前对于橡胶初加工废水污染土壤的危害及修复关注度不够。前期从天然橡胶加工厂附近的受污染土壤中分离和驯化出了一种以芽孢杆菌为优势菌属的高效降解菌群命名为WR-2。WR-2在橡胶初加工废水降解方面表现出优异的性能,但在应用上WR-2需要一定的准备期及环境适应期。因此,本研究采用包埋法,以海藻酸钠–木薯渣生物质炭、甘蔗渣、椰糠3种载体材料及1.5%、2%两种材料浓度下进行固定化小球制备实验,同时进行橡胶初加工废水中典型的胶清废水污染土壤的固定化小球模拟修复实验,研究WR-2的包埋固定化基础性能及其生物修复能力。经过6组固定化微球在机械性能和对污染土壤的修复效果方面比较发现,2%椰糠改良固定化微球不仅具有较优良的机械性能,在模拟修复实验中对污染土壤的有机质（TOC）降解率高达（96.7%）,土壤二硫化四甲基秋兰姆（TMTD）在14 d全部降解,且使土壤pH从4.42调节至中性,土壤阴阳离子总数降低并具有良好的微生物活性。2%椰糠固定化小球极大地改善了土壤微观结构及土壤通气性和排水性。因此,海藻酸钠2%椰糠作为WR-2的固定化小球制备方式,对优化橡胶初加工废水污染土壤的生物修复能力具有效果优和即时应用的作用。