共查询到19条相似文献，搜索用时 187 毫秒
1.
本文在太平洋深海沉积物中分离得到一株孔雀石绿降解菌株,鉴定命名为Tenacibaculum sp.HMG1。通过菌株生长实验和高效液相色谱的研究表明, HMG1菌株可以在20 mg/L的孔雀石绿中维持较快的生长速率,并且在12 h内可降解98.8%的孔雀石绿,这证明该菌株具有很高的孔雀石绿耐受能力和降解活性。通过基因组测序在HMG1菌株发现一条过氧化物酶基因可能参与了孔雀石绿的降解,随后利用原核表达获得了相应的重组蛋白。实验表明,该重组过氧化物酶具有极强的活性,可在1000 mg/L的孔雀石绿中发挥降解功能。本文利用液相色谱-质谱联用（LC-MS）技术对孔雀石绿的菌株降解产物和重组酶降解产物进行鉴定,并基于鉴定结果推测了两种降解途径。结果发现两种降解方式存在共同的降解途径。此外,孔雀石绿降解条件的实验结果证明重组过氧化物酶可以在低温（20℃）、复杂的pH值（6.0–9.0）、高盐度（100 mmol/L）、金属离子和EDTA等反应条件下依旧维持很高的孔雀石绿降解活性。以上实验结果表明,HMG1菌株和重组过氧化物酶均在孔雀石绿污染生物修复方面具有很大潜力。 相似文献
2.
翻译控制肿瘤蛋白Fortilin是1种多功能蛋白,参与重要的细胞活动.并且,对虾fortilin通过抑制病毒复制干扰病毒感染.参照Genbank中fortilin基因序列设计引物,PCR扩增得到凡纳滨对虾fortilin目的基因片段,EcoR I/XbaI双酶切插入表达载体pGAPZαA,转化大肠杆菌,经博来霉素(Zeocin)抗性筛选及测序分析,获得分泌型重组酵母表达载体pGAPZαA-F.酶切线性化后,经电穿孔法转入毕赤酵母细胞X-33,经Zeocin抗性筛选,得到阳性转化子.表达产物的上清液经SDS- PAGE电泳和质谱分析,表明在酵母中成功表达fortilin.以体外原代培养的对虾血细胞检测重组蛋白的免疫活性,结果显示重组蛋白显著提高了血细胞酚氧化酶和超氧化物歧化酶活性.该结果为下一步研究重组fortilin 在对虾养殖中的应用奠定了基础. 相似文献
3.
对具有高淀粉酶活性的南极菌Pseudoalteromonas sp. A211-5的全基因组数据进行了分析和筛选,筛选获得α-淀粉酶疑似序列amy3809,并采用基因工程手段对该基因的功能和性质进行验证和分析。首先,以淀粉酶Amy3809的完整开放阅读框(ORF)为模板设计特异引物,克隆获得amy3809的全序列并对其进行重组表达,获得的重组蛋白采用镍柱进行分离纯化;DNS-还原糖法测定重组酶的酶学性质;薄层层析(TLC)技术对Amy3809的酶解产物进行分析。实验结果:1)克隆获得的amy3809成功地连接到pET-30a载体,并在工程菌E.coli BL21(DE3)中实现了高效表达,纯化的重组酶Amy3809分子量为67 kDa;2)重组酶Amy3809在10~40℃的范围内仍能保持85%以上的酶活,但随着温度的升高酶活迅速降低,70℃时几乎失活,表明该酶具有良好的低温耐受特性及热敏感性;3)最适pH为7.0,在pH 5.0~10.0的范围内仍能保持50%以上的活性;4)金属离子Na+,K+和Ca2+均能提高Amy3809的活性,而Cu2+,Fe2+,Mg2+和EDTA则能显著降低Amy3809的活性;5)Amy3809的酶解产物主要为麦芽四糖、麦芽三糖、麦芽糖和葡萄糖。由此可知,南极菌产的α-淀粉酶Amy3809,具有良好的低温耐受特性,热敏感性和较广的pH耐受范围,并能够有效地将淀粉降解为低聚糖和葡萄糖,因而具有潜在的工业应用前景。 相似文献
4.
海水胞外酶活性可以指示有机物的分布特征以及微生物的营养状况。我们测定了2017年3月25日至4月15日黄海和东海44个大面站以及2018年4月28日至29日胶州湾湾口附近海域10个站位表层海水中的8种胞外酶活性并研究了其分布特征。2017年春季黄、东海表层海水中碱性磷酸酶和脂肪酶活性较高,高值区出现在苏北沿岸和南黄海中部,碱性磷酸酶与磷酸盐浓度之间呈正相关。其余6种酶(肽酶、几丁质酶、纤维素酶、α-D-葡萄糖苷酶、β-D-半乳糖苷酶、木糖苷酶)活性高值区出现在长江口以东的外海,东海的β-D-半乳糖苷酶、木糖苷酶平均酶活性显著高于黄海。8种酶活性平均值排列顺序由大到小为:碱性磷酸酶、脂肪酶、肽酶、几丁质酶、α-D-葡萄糖苷酶、β-D-半乳糖苷酶、纤维素酶、木糖苷酶,其中α-D-葡萄糖苷酶和β-D-半乳糖苷酶的活性基本一致。2018年春季胶州湾附近海域海水中碱性磷酸酶、脂肪酶、木糖苷酶活性分布为近岸高于远岸,几丁质酶活性为近岸低于远岸。8种酶活性平均值排列顺序由大到小为:碱性磷酸酶、脂肪酶、肽酶、木糖苷酶、α-D-葡萄糖苷酶、β-D-半乳糖苷酶、几丁质酶、纤维素酶,其中几丁质酶和纤维素酶的活性基本一致。黄海的碱性磷酸酶和脂肪酶平均酶活性均显著高于东海和胶州湾附近海域。糖类水解酶(几丁质酶、纤维素酶、α-D-葡萄糖苷酶、β-D-半乳糖苷酶、木糖苷酶)平均酶活性在黄海最低。本文的结果对于理解中国近海海水有机碳的分布、浮游植物及异养细菌对有机碳的降解具有重要意义。 相似文献
5.
从来自深海沉积物的太平洋火色杆菌(Flammeovirga pacifica)的基因组中发现了一个全长为1 875 bp的α-淀粉酶基因amy608.该基因在数据库中找不到具有同源性的序列,而所编码的Amy608蛋白与已注册蛋白的氨基酸序列相似性最高仅为56%,但具有α-淀粉酶水解活性所必需的保守基序DXEXD.进化树分析表明其属于糖苷水解酶13(GH13)家族第二亚家族.构建了p ColdΙ-amy608表达载体,在大肠杆菌中进行重组Amy608蛋白的异源表达,并采用镍离子亲和层析柱对重组蛋白进行了纯化.酶学性质分析表明,重组酶Amy608的最适作用温度为40℃,在40℃保温4 d后,仍保留70%以上的酶活,显示出良好的中温热稳定性;最适p H值为7.0,p H值范围在6~9时仍保留60%以上的酶活力,表明该酶具有较宽的p H值作用范围.Ca2+、Na+、K+对α-淀粉酶Amy608有激活作用,尤其是Ca2+可使酶活显著提高40%.薄层色谱分析结果显示该酶水解可溶性淀粉的最终产物以葡萄糖为主,表明该酶是一个糖化型淀粉水解内切酶.这些结果表明Amy608为GH13家族第二亚家族中一个新型的α-淀粉酶. 相似文献
6.
采用PCR扩增、构建重组高效表达载体的方法,进行了牙鲆多聚免疫球蛋白受体(pIgR)基因的克隆、原核表达研究,并利用SDS-PAGE、western-blot及ELISA方法对纯化的重组蛋白特性进行了分析。结果表明,PCR扩增出pIgR开放阅读框(ORF)基因全长为1005 bp,所构建的pET-32(a)-pIgR重组质粒经PCR和双酶切鉴定含有ORF全长基因。SDS-PAGE结果表明,表达的目的蛋白相对分子质量为58 kDa,与理论预期值一致,IPTG诱导6h后表达量趋于稳定,经亲和层析纯化得到高纯度的重组蛋白。Western-blot结果表明,重组pIgR能够与鼠抗His-tag单克隆抗体发生特异性反应;ELISA结果表明重组pIgR能够与牙鲆IgM发生特异性结合。本研究获得了纯化的重组pIgR,证明其具有IgM结合活性,为下一步研究牙鲆pIgR的转运机制及在黏膜免疫防御中的作用机理提供了分子基础。 相似文献
7.
C-型凝集素(C-typelectin,CTL)是甲壳类动物体液免疫中重要的免疫因子之一。但CTL基因在溶藻弧菌对三疣梭子蟹(Portunustrituberculatus)感染的过程中的表达及抗病机制尚有待进一步研究。为初步阐明CTL基因SNP E4-205 C/T位点与抗溶藻弧菌感染相关的分子机制,对不同基因型梭子蟹进行溶藻弧菌感染,通过绝对定量方法分析不同基因型梭子蟹肝胰腺、肌肉组织中溶藻弧菌的复制情况。发现溶藻弧菌感染后12 h内梭子蟹肝胰腺、肌肉组织中C/C组细菌数量显著高于T/T组,结果表明T/T组梭子蟹抗感染能力显著高于C/C组。进一步对该位点非同义突变(ACT-ATT)导致的一个氨基酸改变(Thr-Ile)的两种蛋白进行体外重组表达,并对两种重组蛋白CTL-ATT及CTL-ACT进行活性分析。发现两种重组蛋白对溶藻弧菌生长均具有一定的抑制作用, CTL-ATT的抑菌活性显著高于CTL-ACT。重组蛋白CTL-ATT与PAMPs的结合活性高于CTL-ACT与PAMPs的结合活性,同时两种蛋白与PAMPs和溶藻弧菌的结合活性具有浓度依赖性。结果表明三疣梭子蟹CTL基因参与机... 相似文献
8.
几丁质是贝类贝壳的重要组成成分,在贝壳形成和生物矿化中具有重要作用。本研究通过对厚壳贻贝的贝壳和外套膜边缘三层褶皱进行形态和组织学观察,发现贻贝角质层壳膜是从外褶皱和中褶皱之间的壳膜沟中分泌,而外套膜三层褶皱间组织形态、细胞类型都存在较大差异。此外,本研究还从厚壳贻贝的角质层和贝壳层中提取并鉴定到β-几丁质成分,并构建了外套膜内褶皱（IF）、中褶皱（MF）、外褶皱（OF）的转录组文库,筛查出几丁质合成、降解、结合相关功能的三类几丁质相关基因,对其结构域和表达模式进行进一步分析,发现几丁质合成酶基因以及β氨基己糖苷酶、二-N-乙酰壳二糖酶等几丁质酶基因主要在内、中褶皱表达,而壳三糖苷酶类的几丁质酶基因和几丁质结合蛋白基因则主要在外褶皱表达。在三褶皱差异表达基因的GO富集分析中,进一步发现内、中褶皱主要参与几丁质的生物合成,而外褶皱则主要参与调控几丁质结合、代谢、角质层色素沉着等过程。本研究揭示了外套膜三褶皱在几丁质合成、代谢、结合等功能方面的差异,其中外褶皱可能在厚壳贻贝的贝壳角质层的形成和色素沉着中发挥更重要的作用。本研究有助于了解贻贝贝壳中的几丁质成分以及几丁质相关基因,为软体动物贝... 相似文献
9.
L8和L10是豚鼠气单胞菌(Aeromonas caviae)CB101的两株转座子突变株,突变位置发生在nagA基因,可以组成型表达几丁质酶.我们构建了基因文库,利用Southern杂交从中筛选到一个克隆子包含了三个基因:nagB、nagA和nagC.序列分析显示nagBAC的转录方向相同,而且基因的间隔仅有几个碱基.根据已有报道我们推测NagC是几丁质酶的转录调控蛋白,转座子插入nagA破坏了下游的调控蛋白基因nagC的转录和表达,从而几丁质酶成为组成型表达.我们通过构建NagC缺失突变株、凝胶阻滞迁移以及对NagC氨基酸序列的分析初步证明了NagC属于ROK(repressors,opening reading frames,and kinases)家族的调控蛋白,是chiI的转录阻遏因子. 相似文献
10.
11.
几丁质酶是甲壳动物顺利完成生理性蜕壳的关键功能酶．已有研究表明几丁质酶是一个多基因家族．根据甲壳动物几丁质酶保守序列设计引物,应用反转录PCR方法从凡纳滨对虾（Litopenaeus vannamei）内脏中扩增得到部分几丁质酶编码基因片段,进一步结合RACE法,克隆得到该几丁质酶完整编码序列．生物信息学分析表明其含有信号肽序列、几丁质酶催化中心序列、PEST连接区和几丁质底物结合部位序列．序列比对发现其与中国对虾（ABB85237．1）、斑节对虾（AF157503．1）和日本对虾几丁质酶（BAA12287．1）具有很高的相似度． 相似文献
12.
Reliable methods have been developed for the quantitative estimation of chitin, chitosan and chitin deacetylase in sediment samples, as well as two methods for the estimation of chitinase in these samples. The differing substrates used in the latter two methods have produced distinct ‘depth profiles’ for chitinase activity which suggest the presence of more than one chitinolytic system in the sediments of the Ythan estuary. Studies on the mineralization of chitin in these sediments have revealed a complex system with a number of unexpected features. The rate of chitin degradation was reduced in late summer, possibly due to the heavy seaweed growth which covers the mud flats at this time, although the potential degradative activities of the enzymes chitinase and chitin deacetylase were higher in summer than in winter. The proportional rates of degradation of chitin, chitosan and cellulose were similar in both the absence and presence of seaweed overgrowth, although the apparent rates of degradation of these substrates were higher in the absence of seaweeds. Core samples have indicated that the major input of chitin to the sediment ecosystem occurs via deposition at the surface, while the input of chitosan (the deacetylated polymer) occurs via its production within the sediment, primarily within the upper 5 cm of sediment. The concentrations of both these polymers decreased sharply below an ‘interface’ between aerobic and anoxic sediments. 相似文献
13.
豚鼠气单胞菌CB101在胶体几丁质诱导下产生多种分子量不同的几丁质酶,其中分子量最大的一个几丁质酶Chi1(约91.5kDa)的编码基因已经被克隆.本文报道从CB101发酵液经硫酸铵盐析,DEAE-纤维素柱层析和MonoQ离子交换柱层析,分离纯化出其中的一个分子量约为60KDa的几丁质酶.N端测序的结果表明该酶是Chi1剪缺掉N末端信号肽部分、C末端A区和两个几丁结合区的产物.构建了CB101基因组的cosm id文库,从中筛选到9个产几丁质酶的克隆子,克隆子分析表明它们包含的几丁质酶基因都是chi1.蛋白分离、纯化和基因克隆的实验都暗示CB101的几丁质酶系统只包含一个几丁质酶基因. 相似文献
14.
An agarase produced by a marine bacterium, a bacterial amylase, a fungal cellulase, and an actinomycetal chitinase were examined for their stability and hydrolytic activity at hydrostatic pressures existing in the deep sea. All four enzymes were from barophobic organisms which fail to grow at pressures exceeding 400 to 500 atm, but some observations were made on the hydrolase activity of barophilic bacteria grown at 500 to 1,150 atm. Al-though differing in the rates at which they catalyzed the hydrolysis of agar, starch, cellulose, and chitin respectively, all four enzymes were found to be active at deep-sea pressures and temperatures. These glycoside hydrolases were found to be much more baroduric than most oxidoreductases, transferases, and ligases (synthetases) which have been investigated. The chitinase and-amylase were about as stable at 1,000 atm as at 1 atm. Cellulase was also baroduric and active at 1,000 atm at pH 4.2, its pH optimum at 1 atm. The pressure tolerance of cellulase was much less at higher pH values characteristic of normal sea water, i.e., near pH 8.2. The activity of agarase (synthesized at 1 atm) was retarded by increased pressure, due largely to the inactivation of this enzyme system. However, barophilic bacteria, isolated from the deep sea, synthesize agarases which are active at 1,000 atm as indicated by the hydrolysis of agar.Contribution from the Scripps Institution of Oceanography, University of California, San Diego, La Jolla, Calif. 92037, U.S.A. 相似文献
15.
Analysis of microbiological and chitinase activities relative to the white shrimpPenaeus setiferus, and its chitinoclastic endosymbionts, have demonstrated the relevance of total chitinolytic processes in penaeid biology. Microorganisms may serve as a direct source of nutrients for the animal as well as in the elaboration of extracellularin situ chitinase enzymes. The enzyme produced by the predominant gut bacteria,Beneckea neptuna, is a moderately actively inducible chitinase while the shrimp has an indigenous constitutive chitinase and chitobiase system. Factors of temperature, pH, ion inhibition and reducing sugar ratios have been compared for the bacterial and animal enzymes. This dual enzyme system suggests that metabolic chitin transformation may play a vital role in crustacean metabolism. 相似文献
16.
生物降解甲壳质的研究 总被引：3，自引：0，他引：3
介绍了利用生物 (主要是微生物 )降解甲壳质的一些研究进展 ,着重讨论了在甲壳质降解中起重要作用的几丁质酶的基本性质和分子生物学特征。指出了生物处理海洋资源 ,开发有用活性物质的应用前景。 相似文献
17.
Ellis J. MaginnCrispin T.S. Little Richard J. HerringtonRachel A. Mills 《Marine Geology》2002,181(4):337-356
Hydrothermal vent fauna, particularly vestimentiferan and polychaete worm tubes, are occasionally preserved in the geological record. The early stages of mineralisation are particularly important in defining whether or not preservation will occur, and they are poorly understood. Tube samples of the polychaete worm Alvinella pompejana collected from 13°N on the East Pacific Rise have been studied to identify the processes occurring during early pyrite/marcasite mineralisation. Iron sulphide mineralisation is present within the walls of the organic dwelling tube, and is induced by microbial fauna preserved within the tube micro-layers. Various microorganisms were observed coating the inner tube surfaces, together with 10-100 μm-sized Fe- and Zn-sulphide particles precipitated from vent fluids. The microbes and particulate sulphides become entombed within the tube wall as further layers of organic material are secreted by the worm, during tube-building episodes. This results in a laminated tube structure being formed, composed of alternating layers of tube material and microbial/sulphide-rich interlayers. The microbial/sulphide layers provide a template for further mineralisation and replacement of the microbes with pyrite while degradation of the organic components occurs. The iron monosulphides mackinawite and greigite have been identified as intermediatory phases that occur as precursor minerals during the formation of pyrite. Later marcasite mineralisation is observed to form over some of the pyritised organic layers. Once mineralisation has replaced most of the organic tube material, the structure will then be preserved along with the host sulphide body. These observations enhance our understanding of the mechanisms of fossil pyritisation in fine-scaled organic structures throughout the geological record. 相似文献
18.
研究了地衣芽孢杆菌（Bacillus licheniformis）对尖吻鲈（Lates calcarifer）的生长及消化酶活性的影响.地衣芽孢杆菌制剂（含量为10×108cfu/g）分别以0.0%（对照组）、0.1%、0.2%、0.3%、0.4%、0.5%和0.6%的比例添加到饲料中,投喂初始体重为17.47±0.19 g的尖吻鲈,养殖时间8周.随着地衣芽孢杆菌添加量的增加,尖吻鲈的增重率和特定生长率逐渐增高,在0.5%组达到最大,增重率提高9.77%,特定生长率提高4.56%;同时该组的肥满度也显著高于对照组（p〈0.05）.尖吻鲈的增重率、特定生长率、饲料系数、存活率、脏体比和肝体比在各组间都没有显著差异（p〉0.05）.尖吻鲈的前肠蛋白酶在0.2%组达到最大,并显著高于对照组（p〈0.05）,其他组别及部位的消化酶则低于对照组或差异不显著.结果表明,地衣芽孢杆菌对尖吻鲈的生长有一定促进作用,但此作用与消化酶变化相关不明显. 相似文献
19.
Algae(and their extracts)are increasingly important for pharmaceutical applications due to the diversity of useful compounds they contain.The genus Fucus contains one of the most studied species,Fucus vesiculosus.The species F.ceranoides differs from the others of the genus by presenting longitudinal air-vesicles and a capacity to survive at low salinities.It is an alga that inhabits the Mondego River estuary(Portugal),at the southern limit of its distribution,and can serve as a role model to understand the effect of a salt gradient on the production of bioactive compounds.We assessed the phenolic content and antioxidant activity of different F.ceranoides extracts(e.g.methanolic,aqueous and polysaccharide)prepared from samples harvested from two different zones to evaluate if the adaptation of F.ceranoides to different salinity levels influenced its chemical composition.The antioxidant activity of the extracts was determined using 1,2-diphenyl-picrylhydrazyl(DPPH)and 2.2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)(ABTS)radicals.These assays demonstrated that the methanolic extract of lyophilized F.ceranoides that grew at low salinities was the most bioactive,i.e.DPPH(IC 50=50.39μg/mL)and ABTS(TEAC=2.42).The total phenolic content(Folin-Ciocalteu method)and the methanolic extract of the lyophilized F.ceranoides collected from a low salinity habitat exhibited the highest phenolic content(PGE=49.48μg/mg of lyophilized extract)amongst those sampled.Thin layer chromatography(TLC)and Fourier transform infrared spectroscopy(FTIR)were used for the identification of compounds in the extracts.This characterization allowed confirmation that the various extracts contained almost the same compounds but with notable quantitative differences.Based on these results,we conclude that there were differences in the quantity of the compounds due to the effect of salinity.The drying methods used were also found to have influenced the quality of the extracted compounds. 相似文献