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海藻糖是一种广泛存在于生物体中的非还原性二糖，具有独特的生物学特性，在医药、农业和食品等领域对细胞和生物分子都起到重要的保护作用。乳酸菌作为食品发酵工业中的重要微生物，近年来逐渐成为研究热点。为了进一步探究海藻糖对于乳酸菌的保护机制，本文综述国内外学者关于海藻糖对生物分子的保护机理及其在保护乳酸菌抵御环境胁迫中的应用。海藻糖对生物分子的保护机理存在4 种假说：“水替代”假说、“玻璃态”假说、优先排阻学说和分子间协同作用学说；海藻糖能够对乳酸菌在脱水胁迫、高温胁迫、冷冻胁迫及氧化胁迫等恶劣环境下起到一定的保护作用。本文为今后将海藻糖作为保护剂，提高乳酸菌对不利环境的抗性提供了理论依据。 相似文献
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海藻糖在保护细胞质膜和生物大分子空间结构,维持胞内渗透压增大的过程中发挥重要作用,海藻糖合成酶是海藻糖合成途径中的一个关键酶。通过检索微孢子虫基因组数据库发现,家蚕微孢子虫(Nosema bombycis)基因组中含有4个海藻糖合成酶基因,其中Nbtps1基因的开放阅读框长1 386 bp,为单外显子结构,编码461个氨基酸残基,预测蛋白质分子质量53.7 k D,等电点5.19。Nb TPS1蛋白序列含有1个糖基转移酶结构域和4个潜在的N-糖基化位点,亚细胞定位预测该蛋白质位于细胞质中。多重序列比对分析结果表明,Nb TPS1与东方蜜蜂微孢子虫(Nosema ceranae)、兔脑炎微孢子虫(Encephalitozoon cuniculi)海藻糖合成酶的氨基酸序列相似度分别为59.0%、54.7%,系统发育进化树显示三者的亲缘关系与之相符。构建重组表达载体Nbtps1/p ET-28a(+)并转化大肠埃希菌Rosetta(DE3)菌株,经IPTG诱导表达及亲和纯化后获得最终质量浓度为1.18 mg/m L的重组融合蛋白His-Nb TPS1。用纯化后的融合蛋白制备兔多克隆抗体Anti-Nb TPS1,利用间接ELISA法测定多克隆抗体的效价达1∶25 600,并通过Western blotting检测验证了该多克隆抗体的特异性,为研究Nb TPS1的生物学功能以及家蚕微孢子虫的间接免疫检测奠定了实验基础。 相似文献
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陈进超孙佳静张相鑫唐志如 《动物营养学报》2019,(2):575-583
腺苷酸激活蛋白激酶(AMPK)是一种在真核细胞生物中广泛存在的丝氨酸/苏氨酸蛋白激酶,作为细胞内最重要的能量感受器,AMPK在细胞生长、繁殖、维持机体能量平衡以及细胞代谢过程中发挥重要的调节作用。AMPK活化主要受细胞内一磷酸腺苷/三磷酸腺苷(AMP/ATP)水平及肝激酶B1、钙调素依赖蛋白激酶活性等因素的影响;在动物处于营养缺乏、热应激、氧化应激等环境中,AMPK通路能做出适应性调整,以降低应激环境对动物的负面影响。本文对AMPK的结构、活性调节以及对处于应激环境中的动物能量代谢的调节进行综述,为缓解畜禽各种应激综合征提供理论参考。 相似文献
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腺苷酸激活蛋白激酶信号通路及其在动物应激过程中的调节功能 总被引：1，自引：0，他引：1
腺苷酸激活蛋白激酶(AMPK)是一种在真核细胞生物中广泛存在的丝氨酸/苏氨酸蛋白激酶,作为细胞内最重要的能量感受器,AMPK在细胞生长、繁殖、维持机体能量平衡以及细胞代谢过程中发挥重要的调节作用。AMPK活化主要受细胞内一磷酸腺苷/三磷酸腺苷(AMP/ATP)水平及肝激酶B1、钙调素依赖蛋白激酶活性等因素的影响;在动物处于营养缺乏、热应激、氧化应激等环境中,AMPK通路能做出适应性调整,以降低应激环境对动物的负面影响。本文对AMPK的结构、活性调节以及对处于应激环境中的动物能量代谢的调节进行综述,为缓解畜禽各种应激综合征提供理论参考。 相似文献
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热休克蛋白是机体受到应激原的刺激后所产生的几高度保守的蛋白质,对维持细胞生存和内环境的稳定着重要作用。其中热休克蛋白70是最重要的一种,它大多数生物中含量最多,在细胞应激后生成最显著。休克蛋白具有多种生物学功能,包括分子伴侣功能、参与免疫反应、抗细胞凋亡功能、抗氧化功能、提高细胞的应激耐受性、促进细胞增殖、参与细胞骨架的形成和修复等等, 相似文献
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野桑蚕、家蚕海藻糖酶基因5''侧翼区的克隆与序列分析 总被引：2，自引：0，他引：2
海藻糖是一种双糖,广泛存在于包括细菌、酵母、植物和无脊椎动物在内的生物体内,主要功能是储备生物代谢所需的能源.在昆虫血液中,海藻糖是一种最重要的糖类,具有供给能量和提供合成生物大分子的底物等功能.其功能主要是通过海藻糖酶(Trehalase)的作用来实现的.家蚕海藻糖酶有可溶型和膜固定型两种类型,可溶型主要存在于蛹期中肠,而膜固定型存在于包括卵巢在内的多种组织.这两种类型酶是由同一个基因表达而形成不同的成熟海藻糖酶.海藻糖酶膜固定型存在于向血液面的卵巢膜内,该基因的表达是由滞育激素诱导调控.卵母细胞通过位于膜内的海藻糖酶,将血液中海藻糖降解成葡萄糖,然后吸收到胞内,最后合成糖原,从而导致蚕卵的滞育.从而海藻糖酶基因是从分子水平来解明家蚕滞育机理的一个重要途径.本实验从野桑蚕、家蚕苏@菊×明@虎基因组中分别克隆了野桑蚕和家蚕的海藻糖酶基因的5′侧翼区片段并进行了测序分析,以供进一步的启动子功能研究,现报道如下. 相似文献
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热应激诱导的氧化应激对动物肠道组织的损伤 总被引：1，自引：0，他引：1
热应激是一种常见的非特异性应激,给畜牧业带来较大的损失。肠道组织在热应激作用下易发生缺血缺氧,肠道细胞产生氧化应激,造成细胞凋亡,引起肠道组织损伤。而肠道作为动物机体吸收营养、屏障病原体最为重要的器官,当其受到损伤时将直接影响到动物机体的生长发育及健康状况。本文从热应激诱导肠道细胞产生氧化应激,氧化应激对肠道的损伤,以及热应激诱导细胞凋亡途径等方面,结合国内外近年来研究进展作一综述。 相似文献
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海藻糖是家蚕微孢子虫(Nosema bombycis,Nb)成熟孢子的主要糖类物质之一,海藻糖酶作为海藻糖代谢的主要催化酶,在微孢子虫的发芽及侵染过程中发挥重要作用。通过生物信息学分析发现,家蚕微孢子虫的海藻糖酶具有4个序列相似度较高的编码基因拷贝(Nb Tre1、Nb Tre2、Nb Tre3和Nb Tre4),除了Nb Tre4编码蛋白质的N端有18个氨基酸组成的信号肽,其他拷贝的编码蛋白质均没有信号肽结构域,但都具有少数的N-糖基化位点,而没有O-糖基化位点,此外,丝氨酸磷酸化位点的比率也较高,二级结构较为简单,仅有螺旋区和低复杂度区。基于生物信息学分析结果,设计特异性引物克隆了Nb Tre1基因,该基因片段长933 bp,编码310个氨基酸,预测蛋白质分子质量36.7 k D,蛋白质的氨基酸序列有2个潜在的N-糖基化位点和14个磷酸化位点。通过构建重组表达载体并转化Escherichia coli Rosetta(DE3)感受态细胞,获得Nb Tre1蛋白的表达菌株,利用IPTG诱导获得大量以包涵体形式表达的融合Nb Tre1蛋白,其分子质量与预期值一致。融合Nb Tre1蛋白经亲和层析纯化后免疫新西兰白兔制备多克隆抗体,利用间接ELISA法测定多克隆抗体的效价达1∶25 600。通过Western blot检测验证了该多克隆抗体的特异性,制备的多克隆抗体能够应用于家蚕微孢子虫海藻糖酶的检测等。 相似文献
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M B Rhodes C A Klucas M L Frey G A Anderson 《Journal of veterinary diagnostic investigation》1989,1(4):324-328
A blocking enzyme-linked immunosorbent assay (ELISA) has been adapted to detect specific antibodies in bovine sera to respiratory syncytial virus using a horseradish peroxidase-labeled monoclonal antibody to the fusion protein of the virus. This assay plus an indirect blocking ELISA and indirect ELISA were used to detect antibodies to the bovine respiratory syncytial virus (BRSV) in 159 field-origin bovine sera. Results of these assays were compared with serum antibody titers measured by the serum neutralization (SN) test. Over a 56-day period, the mean neutralization titers and the mean delta absorbance values for the blocking ELISA, on the same sera, showed similar declines. However, the calculated correlation coefficients between mean SN titer and mean absorbance value for the blocking ELISA of the individual sera ranged from -0.2 to -0.5 depending on the source of sera. Similar values were obtained whether using crude or purified viral antigen in the assays. Corresponding calculated correlation coefficients were generally higher for the indirect blocking ELISA or indirect ELISA than for the blocking ELISA. The blocking ELISA was between 70 and 64% as sensitive as the serum neutralization test with a specificity of 100 or 90% using the crude and purified viral antigen, respectively. The indirect blocking ELISA and indirect ELISA had similar calculated sensitivities and specificities. The blocking ELISA was faster to run than either of the other ELISA's or the neutralization test. Further, nonspecific background absorbance was obviated because the blocking ELISA detects antibodies to 1 specific viral protein, the fusion protein. These studies suggest that the blocking ELISA should be useful as a serological test for BRSV antibodies. 相似文献
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Mycoplasma ovipneumoniae NCTC 10151(T) and four new isolates from UK sheep flocks were compared. Only glucose and pyruvate were used as energy sources by the five strains: glucose was the best energy source for the type strain, pyruvate supported better growth of the new strains. Whole cell protein patterns and antigenic profiles showed high similarity between all five strains. The new isolates fell into two groups in ELISA tests. Serum samples from 30 pneumonic sheep were assessed for M. ovipneumoniae infection and Mycoplasma arginini co-infection. Fourteen (out of 30) serum samples were positive for M. ovipneumoniae both by ELISA and immunoblotting. Twelve antigenic proteins of M. ovipneumoniae were detected in infected serum samples: the antigen patterns were unique, with between one and at least seven occurring in any one sample. All serum samples were designated as negative for M. arginini antibodies by both ELISA and immunoblotting. 相似文献
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For the surveillance of trichinellosis, the digestion method is reliable but also labour intensive. The serological methods for the detection of Trichinella-specific antibodies using ELISA offer a sensitive and relatively specific alternative. For serological studies, sera or plasma from blood samples are the most common source of antibodies, but although the concentration of antibodies is approximately 10-fold lower, muscle fluid can be a good alternative particularly for testing of wildlife samples. In the present study, an indirect ELISA technique was evaluated on both sera and muscle fluids from experimentally infected foxes, pigs, and wild boars using both excretory/secretory (E/S) antigens and a synthetic glycan antigen, beta-tyvelose. Although the synthetic antigen appears to be less sensitive than the E/S antigens, Trichinella-specific IgG antibodies were detected in both serum samples and muscle fluid samples from pigs, wild boars and foxes infected at levels which would be important for food safety or represent a significant reservoir for further transmission. 相似文献
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K Sestak Z Zhou D I Shoup L J Saif 《Journal of veterinary diagnostic investigation》1999,11(3):205-214
The spike (S) glycoprotein of the Miller strain of transmissible gastroenteritis virus (TGEV) was recently cloned and expressed in baculovirus. The recombinant S protein was used as the coating antigen in a competition (blocking) enzyme-linked immunosorbent assay (ELISA) in combination with monoclonal antibodies to the S protein epitope A (conserved on TGEV and porcine respiratory coronavirus [PRCV]) or epitope D (present on TGEV only) to differentiate PRCV- from TGEV-induced antibodies. One set (set A) of 125 serum samples were collected at different times after inoculation of caesarean-derived, colostrum-deprived (n = 52) and conventional young pigs (n = 73) with 1 of the 2 porcine coronaviruses or uninoculated negative controls (TGEV/PRCV/negative = 75/30/20). A second set (set B) of 63 serum samples originated from adult sows inoculated with PRCV and the recombinant TGEV S protein or with mock-protein control and then exposed to virulent TGEV after challenge of their litters. Sera from set A were used to assess the accuracy indicators (sensitivity, specificity, accuracy) of the fixed-cell blocking ELISA, which uses swine testicular cells infected with the M6 strain of TGEV as the antigen source (ELISA 1) and the newly developed ELISA based on the recombinant S protein as antigen (ELISA 2). The sera from set B (adults) were tested for comparison. The plaque reduction virus neutralization test was used as a confirmatory test for the presence of antibodies to TGEV/PRCV in the test sera. The accuracy indicators for both ELISAs suggest that differential diagnosis can be of practical use at least 3 weeks after inoculation by testing the dual (acute/convalescent) samples from each individual in conjunction with another confirmatory (virus neutralization) antibody assay to provide valid and complete differentiation information. Moreover, whereas ELISA 1 had 10-20% false positive results to epitope D for PRCV-infected pigs (set A samples), no false-positive results to epitope D occurred using ELISA 2, indicating its greater specificity. The progression of seroresponses to the TGEV S protein epitopes A or D, as measured by the 2 ELISAs, was similar for both sets (A and B) of samples. Differentiation between TGEV and PRCV antibodies (based on seroresponses to epitope D) was consistently measured after the third week of inoculation. 相似文献
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Silvia M Estein Pablo C Baldi Raúl A Bowden 《Journal of veterinary diagnostic investigation》2002,14(5):407-411
The aim of this work was to compare the performance of 6 serological tests using outer or internal antigens from Brucella for the diagnosis of Brucella ovis infection in sheep in an endemic area. Outer membrane antigens included a hot saline extract (HS) and the rough lipopolysaccharide (R-LPS) from B. ovis. Internal antigens were LPS-free total cytosolic proteins (CP) and an 18-kDa cytosolic protein (p18) from Brucella spp. Sera from 200 sheep from naturally infected flocks were assayed by agar gel immunodiffusion test (AGID) and by complement fixation test (CFT), both using HS, and by 4 ELISA using HS, R-LPS, CP, and p18, respectively. The percentage of positive results was 45.5% for ELISA with HS, 42.0% for ELISA with p18, 39.5% for CFT, 33.5% for ELISA with R-LPS, 29.0% for ELISA with CP, and 18.0% for AGID. Taking CFT as the reference test for calculating relative test parameters, the ELISA with HS had the best sensitivity (96.2%), while AGID and the ELISA with R-LPS had the best specificity (96.6%). The ELISA with CP was not more sensitive than the ELISA with p18 (67.1% vs. 79.7%) in spite of the higher number of antigens in CP. The lower relative sensitivity of tests using internal antigens might reflect a lack of antibodies to cytosolic proteins in some infected animals or a shorter persistence of these antibodies relative to antibodies to outer membrane components after recovery from infection. 相似文献
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A blocking ELISA was developed to detect antibodies directed against porcine epidemic diarrhea virus (PEDV). The PEDV antigen was first incubated with dilutions of test sera. Any antigen that was not blocked by antibodies in the serum was assayed in a double-antibody sandwich ELISA, using 2 monoclonal antibodies directed against different antigenic sites on PEDV as capture and detecting antibodies, respectively. The blocking ELISA was compared with a fixed-cell ELISA that used monolayers of Vero cells infected with PEDV prototype strain CV777 as a solid phase and a conjugate of an IgG-specific monoclonal antibody for antibody detection. Pigs were inoculated with PEDV strain CV777 or 1 of 2 field isolates, and antibody responses were measured by use of the 2 tests. Antibodies were detected by the blocking ELISA as early as postinoculation day 7 and, by the fixed-cell ELISA, as early as postinoculation day 14. From day 14 on, antibody titers for both tests correlated highly. Titers for the fixed-cell ELISA were 5.4 times higher than those for the blocking ELISA. The latter technique is easier to perform and discriminates well between infected and noninfected pigs, which makes this test useful for routine diagnosis and serologic surveys of porcine epidemic diarrhea. 相似文献
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An indirect enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to maedi-visna virus (mvv) in sheep is described, in which microtitre plates are with a partly purified preparation of mvv. The antibodies bound are detected by a horseradish peroxidase conjugate.The results obtained with ELISA on a total of 493 serum samples from several commercial flocks were compared to those of a routine agar gel precipitation test (AGPT) and a complement fixation test (CFT).All samples which scored positive in AGPT, CFT or both (20.8%) were also found positive by ELISA. In addition, with ELISA a further 11.5% of the samples were positive. Serum samples from maedi-free flocks, from sheep suffering from sheep pulmonary adenomatosis and from lambs immunized against other viruses were all negative by ELISA. The assay has been used routinely for some years and proved to be specific, sensitive and suited for screening of large numbers of serum samples. 相似文献
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Enzyme-linked immunosorbent assay for detecting antibodies to Brucella abortus in bovine milk and serum 总被引：1，自引：0，他引：1
F C Heck J D Williams J Pruett R Sanders D L Zink 《American journal of veterinary research》1980,41(12):2082-2084
An enzyme-linked immunosorbent assay (ELISA) method was evaluated for the detection of antibodies to Brucella abortus in cows milk. Milk samples from seropositive or -negative cows were sed to determine the distribution of absorbance values to classify milk as ELISA positive or ELISA negative. Brucella abortus was isolated from milk samples from 10 (45%) of the 22 cows whose milk and serum were ELISA positive. The ELISA was evaluated and determined to be an appropriate method for detecting antibodies to B abortus in bovine milk. 相似文献