橡胶树半胱氨酸蛋白酶基因HbCP3的克隆与表达特性分析

半胱氨酸蛋白酶是一类重要蛋白水解酶,其广泛参与植物的生长发育及各种生物与非生物胁迫的应答。根据EST序列信息设计引物,从橡胶树的根中分离到1条1 056 bp的c DNA。该序列包含1个1 023 bp的开放阅读框(ORF),13 bp的5'UTR和20 bp的3'UTR。ORF预测编码340个氨基酸,相对分子质量为37 350,等电点为5.24,属于液泡定位的分泌型蛋白。该蛋白含有1个Inhibitor_I29和1个Peptidase_C1A结构域,可归为木瓜蛋白酶C1家族。该蛋白与蓖麻(Ricinus communis)和杨树(Populus trichocarpa)中同源蛋白的相似性都在80%以上,将基因命名为Hb CP3。表达分析显示,在橡胶树的根、树皮、木质部、芽、叶片、叶柄和乳管等组织中,基因仅在根中表达。     

Colostral Trypsin-Inhibitor Capacity in Different Animal Species

Colostral trypsin-inhibitor capacity was monitored during the first two weeks from parturition. The colostrum of the mare, sow, cow and ewe showed high antitrypsin activity at parturition, decreasing to about one hundredth during the first week. Canine milk remained on a relatively high antitrypsin level, and human milk was poor in antitrypsin from childbirth. The antitrypsin content seems to parallel the known changes in the colostral immunoglobulin levels of different species. The role of antitrypsin in protection of immunoglobulins from proteolytic damage during passive transfer of immunity to the newborn is obvious.     

Protein degradation and protease activity of chum salmon (Oncorhynchus keta) muscle during spawning migration

Sarcoplasmic protein content decreased significantly and myosin heavy chain was degraded gradually during spawning migration of chum salmon (Oncorhynchus keta). Acid and neutral proteinase activities increased significantly during spawning migration. These proteinase activities were higher in females than in males. High levels of acid proteinase were not caused by the injury of lysosomal membranes. Considering the physiological states of chum salmon, neutral proteinase activity might be related to the degradation of muscle protein. As the changes in serum sex steroids were similar to those in the protease activities during spawning migration, it was considered that high levels of protease activity during spawning migration were closely related to the serum levels of androgens.     

Co~（2＋）对渗透胁迫下小麦幼苗叶片乙烯产生率及蛋白酶活性的影响

在－１．０ＭＰａＰＥＧ渗透胁迫下，小麦幼苗叶片乙烯产生速率和蛋白酶活性随胁迫时间延长递增，蛋白质和叶绿素含量递减。在－１．０ＭＰａＰＥＧ溶液中加入２ｍｍｏｌ／ＬＣｏＣｌ２处理可以抑制乙烯的产生及蛋白酶的活性，对蛋白质和叶绿素起到保护作用。同时还观察到，渗透胁迫过程中乙烯的产生速率与叶片的衰老过程具有正相关性，加入Ｃｏ２＋能一定程度的减轻渗透胁迫造成的伤害。     

Experimental Bubalian Fasciolosis: Kinetics of Antibody Response using 28 kDa Fasciola gigantica Cysteine Proteinase as Antigen

Tropical Animal Health and Production -     

枯草芽孢杆菌发酵鱼粉产蛋白酶及其酶解效果

为开发以鱼粉为原料的优质饲料蛋白源以及相关保健食品,开展了用不同鱼粉为氮源发酵枯草芽孢杆菌生产蛋白酶及其酶解效果的研究,比较了不同菌种的发酵效果,确定了最佳发酵时间.通过蛋白酶活性测定和氨基态氮含量测定分析了不同菌种的产酶能力和酶活性大小及其酶解效果.结果表明,用罗非鱼鱼粉和处理过的鳕鱼鱼粉发酵商业化的工业菌种枯草芽孢杆菌产生的酶活性显著高于鲢鱼鱼粉（P〈0.05）,发酵后上清液中的氨基态氮含量也显著高于鲢鱼鱼粉（P〈0.05）,即工业化的枯草芽孢杆菌对不同鱼粉的发酵效果不同,罗非鱼鱼粉和处理过的鳕鱼鱼粉发酵效果显著优于鲢鱼鱼粉.     

月季花瓣丝氨酸蛋白酶基因RhSep1的克隆及生物信息学分析

利用RT-PCR技术,从切花月季品种"Samantha"中克隆丝氨酸蛋白酶基因RhSep1。利用生物信息学技术对所得到的丝氨酸蛋白酶RhSep1序列进行结构与功能预测。结果表明:RhSep1基因全长2 442 bp,开放阅读框编码769个氨基酸,推定该丝氨酸蛋白酶分子质量为80.48 kD。通过NCBI和MEROPS肽酶数据库等对Rh-Sep1进行Protein Blast,发现RhSep1具有肽酶S8₃家族SA的典型结构域Petidase_S8₃（序列为Tyr-108-Leu-341）和PA_subtilisin_like的结构域（序列为Tyr-348-Ile-474）。预测RhSep1可能具有信号肽、明显疏水区和典型跨膜区,同时RhSep1氨基酸序列中存在较多蛋白激酶C和酪蛋白激酶Ⅱ磷酸化位点、豆蔻酰化位点。这些活性位点往往与蛋白的磷酸化、G蛋白相互作用等信号转导事件有关,RhSep1可能存在复杂的蛋白水平调控机制。     

斑点叉尾鮰鱼肠蛋白酶的分离纯化及其酶学性质

以斑点叉尾鮰Ictalurus punctatus鱼肠道为原料提取蛋白酶,对分离条件进行了优化,并对部分酶学性质进行了研究。结果表明：鱼肠内的蛋白酶在硫酸铵饱和度为70%时,所得沉淀中酶活力最高。经过阴离子交换层析分离后,蛋白酶的纯化倍数达到了218倍。该蛋白酶的最适温度为55℃,最适pH为7.5,具有良好的低温稳定性和酸碱稳定性,米氏常数为5.4 g/L。K^＋和Ca^2＋离子对该蛋白酶活性有较弱的激活作用,Mg^2＋则能显著激活蛋白酶活性;Na＋和Zn2＋对蛋白酶的活性有较弱抑制作用,而Cu^2＋和EDTA能显著抑制蛋白酶活性。     

桔小实蝇丝氨酸蛋白酶基因（BdorSer）的克隆与表达分析

【目的】克隆桔小实蝇丝氨酸蛋白酶基因（Bactrocera dorsalis serine protease,BdorSer）,研究BdorSer在不同组织和不同发育时期的表达情况,探索其可能参与的生理过程。【方法】采用反转录聚合酶链式反应(RT-PCR)和快速扩增cDNA末端(RACE)技术克隆BdorSer,对其编码蛋白氨基酸序列信息和进化关系进行分析;采用实时荧光定量PCR（real-time PCR）方法,研究BdorSer mRNA在桔小实蝇不同组织及不同发育时期的相对表达量。【结果】克隆获得了BdorSer,该基因ORF全长1 239 bp。氨基酸序列一致性分析表明,BdorSer与拟暗果蝇（Drosophila pseudoobscura）丝氨酸蛋白酶（登录号XP_001352960）氨基酸序列的一致性最高,为65.8%。实时荧光定量PCR分析表明,BdorSer mRNA在桔小实蝇雌雄虫的不同组织中都有表达,且在触角中的表达量最高,分别是雌虫胸部的43.6和26.8倍。BdorSer mRNA在生殖节中表现出较大的性别差异表达特征,雄虫生殖节中的mRNA表达量是雌虫的18.25倍。BdorSer mRNA几乎在昆虫发育的各个时期都有表达,其中在卵及1,2,3龄幼虫中的表达量分别是10 d 蛹的 0.68,1.63,4.31和15.8倍。BdorSer mRNA随蛹的发育其表达量逐渐减低,1,4,7 d蛹中的表达量分别是10 d蛹的325,125和41倍。【结论】克隆获得了BdorSer,其表达的BdorSer蛋白可能在雄虫的生殖生理中发挥了作用,同时也可能参与了桔小实蝇的变态发育过程,尤其是1 d蛹的发育过程。     

Brodifacoum is effective against Norway rats (Rattus norvegicus) in a tyrosine139cysteine focus of anticoagulant resistance in Westphalia,Germany

BACKGROUND: The tyrosine to cysteine amino acid substitution at location 139 of the vkorc1 protein (i.e. tyrosine139cysteine or Y139C) is the most widespread anticoagulant resistance mutation in Norway rats (Rattus norvegicus Berk.) in Europe. Field trials were conducted to determine the incidence of the Y139C mutation at two rat‐infested farms in Westphalia, Germany, and to estimate the practical efficacy against them of applications, using a pulsed baiting treatment regime, of a proprietary bait (Klerat^?) containing 0.005% brodifacoum. RESULTS: DNA analysis for the Y139C mutation showed that resistant rats were prevalent at the two farms, with an incidence of 80.0 and 78.6% respectively. Applications of brodifacoum bait achieved results of 99.2 and 100.0% control at the two farms, when measured by census baiting, although the treatment was somewhat prolonged at one site, possibly owing to the abundance of attractive alternative food. CONCLUSION: The study showed that 0.005% brodifacoum bait is fully effective against Norway rats possessing the Y139C mutation at the Münsterland focus and is likely to be so elsewhere in Europe where this mutation is found. The pulsed baiting regime reduced to relatively low levels the quantity of bait required to control these two substantial resistant Norway rat infestations. Previous studies had shown much larger quantities of bromadiolone and difenacoum baits used in largely ineffective treatments against Y139C resistant rats in the Münsterland. These results should be considered when making decisions about the use of anticoagulants against resistant Norway rats and their potential environmental impacts. Copyright © 2012 Society of Chemical Industry