草鱼肠道-菌株产纤维素酶的分离纯化及性质研究

从草鱼肠道分离出1株枯草芽孢杆菌,有较强的活性.该菌产生的纤维素酶粗酶液,经盐析、透析,并通过葡聚糖凝胶层析柱Sephadex G-75分离纯化,经SDS-PAGE后表明第2峰已纯化,得到一种相对分子质量约为62.43 kD纤维素酶.分离纯化后该酶的比活力提高了2.924倍,回收率为6.38%.酶学试验研究表明:该酶的最适反应温度为55℃,最适pH值为7.0;Lineweaver-Burk法求得动力学参数,Km和Bmax分别为1.02×10-3g/mL、2.727×10-2mg/(mL·min).     

皱纹盘鲍内脏酶的酶学性质及褐藻胶裂解酶的分离纯化

采用(NH4)2SO4分段盐析、透析、阴离子(DEAE-52)交换柱层析、SephadexG-200凝胶柱层析等分离纯化技术，通过SDS-PAGE电泳分析了皱纹盘鲍内脏酶的组成，结果表明鲍内脏酶主要含有两种褐藻胶裂解酶Ⅰ, Ⅱ，一种纤维素酶，一种琼脂酶。对酶的酶学性质分析结果表明两种褐藻胶裂解酶Ⅰ, Ⅱ的最适pH分别为8.6, 7.2，最适温度为35 ℃，分子量分别为35.2 ku, 67 ku；两种褐藻胶裂解酶的热稳定性比较差，且易受金属离子影响；纤维素酶的最适pH为5.0，最适温度为40 ℃。并确定了皱纹盘鲍内脏酶分离纯化的方法及参数，为进一步研究鲍内脏复合酶的性能提供了基础参数。图14表3参12 关键词：皱纹盘鲍； 褐藻胶裂解酶； 纤维素酶； 纯化 E-mail：wqk320@dlfu.edu.cn     

1种致病性鳗弧菌胞外蛋白酶的分离纯化及性质

从山东省莱州海区自然发病的花鲈（Lateolobrax japanicus）体内分离到1株致病性鳗弧菌，经硫酸铵盐析、DEAE-Sepharose Fast Flow和Sephadex-G100凝胶层析等方法从其培养液中分离纯化了1种胞外蛋白酶。用SDS-PAGE电泳测得蛋白质的分子量为36.7kD，最适温度为50℃，对热不稳定，70℃15min完全失去活力；最适pH为7.0；1mmol/L的PMSF对酶活性无影响，部分金属离子如Cu^2 、Fe^2 、Fe^3 、Zn^2 对酶活力有抑制作用，而Ca^2 对酶有一定程度的激活作用，1mmol/L EDTA能完全抑制酶的活性，表明该酶是1种金属蛋白酶。     

饲用植酸酶和纤维素酶最佳酶解条件的研究

采取正交试验的方法，研究植酸酶和纤维素酶对配合饲料的酶解效果以及酶解的最适条件。结果表明：饲料经植酸酶酶解后，无机磷含量增加了6．1％～47．2％，平均增加了25．9％；植酸酶酶解的最适条件为：55℃，pH4．6，作用时间为80分钟。饲料经纤维素酶作用后，葡萄糖含量增加了36．7％-80．7％，平均增加了54．8％；在50℃，pH5．0，作用时间为80分钟时，纤维素酶的酶解效果最好。     

大菱鲆消化道蛋白酶的初步研究

以-龄大菱鲆为材料,研究养殖水温下不同pH条件胃、幽门盲囊、前肠、中肠、后肠中蛋白酶的活性,确定饵料中蛋白质的主要消化部位为胃和前肠,各部位主要蛋白酶的最适作用pH依次为2.0、8.0、8.0、8.5、8.0;蛋白酶单位酶活力从高到低依次为幽门盲囊＞胃＞前肠＞中肠＞后肠.以大菱鲆胃为材料,经Tris-HCl缓冲液抽提、硫酸铵分级分离,DEAE-Sepharose Fast Flow离子交换层析、Sephadex G-100凝胶过滤层析分离纯化,获得大菱鲆胃蛋白酶的电泳纯制品,并对该酶的性质进行了研究.结果显示:纯酶的分子量为39900Dal,纯酶最适反应pH为2.0,最适反应温度40℃;pH稳定范围1.0-9.0,40℃以下酶活性稳定;Mn2+和Cu2+可激活大菱鲆胃蛋白酶,Fe3+对酶活性有抑制作用.     

凡纳滨对虾蛋白酶的分离纯化及生化特性

采用Tris-HCl缓冲液抽提、Sephadex G-100凝胶过滤层析、DEAE-Sepharose F.F.阴离子交换层析和SDS-PAGE等方法,分离和纯化了凡纳滨对虾蛋白酶,研究了其生化特性.试验结果表明,该蛋白酶粗提物经层析后,得到聚丙烯酰胺凝胶电泳纯的一酶组分.该蛋白酶的比活力从128.35 U/mg增至1835.65 U/mg,提高了14.3倍,产率达31.9%.SDS-PAGE显示该蛋白酶只含一条谱带,相对分子量为25 kD.以酪蛋白为底物,该酶的最适pH为7.0,最适温度为60 ℃, 在50 ℃以下,比较稳定,放置1 h后活性仍超过60%,而超过50 ℃时蛋白酶活性急剧下降,60 ℃放置1 h后,酶活性只残留4.7%.10 mmol/L EDTA、Fe~(2+)、Ba~(2+)和Zn~(2+)对该蛋白酶有较强的抑制作用,抑制率分别为84%、53%、43%和38%,10 mmol/L Mg~(2+)和Cu~(2+)对蛋白酶活性有轻微抑制作用,而10 mmol/L Ca2+能显著促进蛋白酶活性,据此推测凡纳滨对虾蛋白酶可能为一种金属蛋白酶.     

大菱鲆肠道蛋白酶的分离纯化及性质的初步研究

以大菱鲆肠道为材料,对大菱鲆消化生理中起重要作用的肠蛋白酶的分离纯化条件和理化性质进行了研究。经Tris-HCl缓冲液抽提,硫酸铵分级分离,DEAE-Sepharose Fast Flow离子交换层析、Sephadex G-100凝胶过滤层析分离纯化,获得大菱鲆肠蛋白酶Ⅰ的电泳纯制品,并对该酶的性质进行了研究。结果显示:纯酶的分子量约为58kD。纯酶最适反应pH为9.0,最适反应温度50℃;pH稳定范围6.0~11.0,30℃以下酶活性稳定;Mn2 可激活大菱鲆肠蛋白酶Ⅰ,Cu2 、Zn2 、Ag 、Fe3 则对酶活性有抑制作用;巯基蛋白酶抑制剂显著抑制大菱鲆肠蛋白酶Ⅰ的活性,金属蛋白酶抑制剂对大菱鲆肠蛋白酶Ⅰ无影响。50℃、pH9.0条件下以双倒数作图法得大菱鲆肠蛋白酶Ⅰ催化酪蛋白水解的Km值为2.92g.L-1。     

卡拉胶降解菌M－2的筛选与产酶性质

从多种海藻表面分离筛选到具有较高卡拉胶降解活性的菌株M-2,对该菌株所产卡拉胶降解酶的部分酶学性质和酶解产物进行分析。研究表明,M-2所产卡拉胶降解酶为诱导酶,具有热不稳定性,反应的最适温度为32～38℃,而最适pH为6～8;酶的反应活性受多种离子的影响。经基体辅助激光解吸电离———飞行时间质谱测试,酶解卡拉胶终产物以四糖和六糖为主,该酶为κ-卡拉胶酶。     

罗非鱼肌肉乙酰胆碱酯酶的纯化、性质及农药敏感性研究

以罗非鱼脑组织为材料,研究了乙酰胆碱酯酶的分离纯化方法、性质及杀虫剂对酶的抑制作用。结果表明,当脑组织的TritonX-100抽提物采用(NH4)2SO4分级盐析、Sephadex G-100凝胶过滤和DEAE-纤维素DE23离子交换柱层析的分离纯化步骤后,可获得纯化的乙酰胆碱酯酶,酶的比活力达到20.628U.mg-1, 纯化倍数139,回收率22.1%。酶作用最适温度为35～40℃,最适pH为7.5～8.0,对碘化硫代乙酰胆碱的Km值为0.183mmol.L-1,有过量底物抑制现象,最佳底物浓度为6mmol.L-1。被测4种农药中,有机磷类杀虫剂对罗非鱼脑AChE的抑制能力高于氨基甲酸酯类,抑制能力分别为：敌敌畏 > 辛硫磷 > 三唑磷 > 灭多威, 4种杀虫剂对罗非鱼脑AChE的I50值均小于5μg.mL-1。     

金乌贼肌肉中三甲胺脱甲基酶的分离纯化及酶学性质

为了提取纯化金乌贼肌肉中的三甲胺脱甲基酶(TMAOase),本研究采用含有0.1 mol/L NaCl、pH 7.0、浓度为20 mmol/L的三羟甲基氨基甲烷(Tris)-醋酸缓冲液提取粗酶液,经透析、浓缩处理后,通过DEAE-52阴离子交换柱层析和Sephacryl S-300柱层析得到了纯化的TMAOase,并对其酶学性质进行了研究。结果显示,经Sephacryl S-300柱层析的TMAOase相比粗酶纯化了209.54倍;粗酶和纯化酶的最适温度分别为55和50°C,当温度高于最适温度时,酶活性开始出现显著下降,粗酶在80°C仍残留21.9%的活性;而纯化酶在80°C时,几乎检测不到酶活;粗酶和纯化酶的最适p H均为7.0,中性条件下表现稳定,在酸性和碱性条件下稳定性下降,p H为9.0时,粗酶残留60.7%的活性,而纯化酶的活性仅为20.5%。以双倒数作图法(Lineweaver-Burk法)测得纯化的TMAOase的Km值为22.8 mmol/L;经SDS-PAGE电泳分析,测得其分子量为21.3 ku;在化学物质中,柠檬酸和CaCl_2对酶活性具有显著的促进作用,H_2O_2和Na_2S对TMAOase活性有显著抑制作用。     

六边形开口方形人工鱼礁阻力系数数值模拟与模型试验比较研究

为了对4种不同开口大小的六边形开口方形人工鱼礁在4种迎流角度下数值模拟和水槽模型试验的阻力系数进行对比验证,分别利用水槽模型和数值模拟试验方法对4种不同开口比（γ_ty=0.0625,0.14,0.25,0.39）六边形开口方形人工鱼礁在4种迎流角度（θ=0°,15°,30°,45°）状态下的阻力进行测定,并计算两种方法的阻力系数。结果表明：（1）在数值模拟和水槽模型试验中,人工鱼礁模型阻力均随着开口比的增大而减小;礁体迎流角度的变化可改变礁体阻力,且在4种迎流角度下,人工鱼礁阻力在θ=30°时最大。（2）人工鱼礁数值模拟与水槽模型试验中,当θ为15°、30°和45°时,阻力系数均随着开口比的增加而增加,具有明显的线性关系,且阻力系数在迎流角度θ=30°时最大。（3）数值模拟与模型试验阻力的相对误差在0.12%~17.18%,平均误差7.43%;礁体阻力系数的相对误差在0.03%~14.64%,平均误差5.26%。阻力及阻力系数误差均在20%以下。水槽模型试验与数值模拟阻力和阻力系数相关系数R分别为0.99和0.80,P<0.001,具有极强的相关性。因此,利用数值模拟精细化研究人工鱼礁水动力性能是可行的。     

Trophic relationships between a native and a nonnative predator in a system of natural lakes

Abstract – Bull trout, a species of char listed as threatened under the US Endangered Species Act, have been displaced from portions of their historic range following the introduction of nonnative lake trout. It has been suggested that competitive exclusion as a result of trophic overlap between bull trout and lake trout may be the causal mechanism associated with displacement of bull trout. This study used stable isotope data to evaluate trophic relationships among native bull trout, nonnative lake trout and other fishes in seven lakes in Glacier National Park (GNP), Montana. Bull trout and lake trout had greater δ¹⁵N values relative to other fishes among lakes (δ¹⁵N ≥ 3.0‰). Lake trout had greater δ¹⁵N values relative to bull trout (δ¹⁵N = +1.0‰). Bull trout had greater δ¹³C values relative to lake trout in six of the seven lakes examined. Although both bull trout and lake trout had greater δ¹⁵N values relative to other fishes within lakes in GNP, differences in δ¹⁵N and δ¹³C between bull trout and lake trout suggest that they are consuming different prey species or similar prey species in different proportions. Therefore, displacement of bull trout as a direct result of complete overlap in food resource use is not anticipated unless diet shifts occur or food resources become limiting. Additionally, future studies should evaluate food habits to identify important prey species and sources of partial dietary overlap between bull trout and lake trout.     

Characterization of a cDNA encoding a homolog of actin-related protein 4 from a marine red alga Porphyra yezoensis

ABSTRACT:   A cDNA ( PyARP4 ) containing an open reading frame for a protein of 573 amino acids was identified in the marine red alga Porphyra yezoensis . The conceptual PyARP4 protein exhibits significant similarity to actin-related protein (ARP) 4 in the terrestrial plant Arabidopsis . Comparison of the deduced amino acid sequence showed moderate sequence identity (30%) to a conventional actin in P. yezoensis , as seen in comparisons between ARP and conventional actins of other organisms. A putative bipartite nuclear localization signal and an actin motif were found within the PyARP4 amino acid sequence. In a phylogenetic analysis, the PyARP4 was found to cluster with the ARP4 of other organisms. The expression level of PyARP4 did not change significantly among four developmental stages of life cycle and was lower than that of a conventional actin. This cDNA therefore may serve as a useful internal standard in gene expression analyses of differentially expressed genes in P. yezoensis .     

Habitat characteristics affecting occurrence of a fluvial species in a watershed altered by a large reservoir

Flooding river valleys following construction of dams restrict fluvial environments to reaches that were formerly headwaters. Whether remaining habitat is suitable for all life stages of fluvial species is poorly understood. A fluvial species, Arctic grayling Thymallus arcticus, showed a dramatic decline following flooding of the Upper Peace River and the formation of the Williston Reservoir. We related landscape and field site‐specific features with occurrence of juvenile Arctic grayling using an information theoretic approach. For the landscape model, an association was identified between stream order and Arctic grayling occurrence although stream order alone was a poor predictor. A positive association between juvenile Arctic grayling occurrence and distance from the Williston reservoir and stream order, as well as a negative association with water temperature and temperature variance, was deemed important for the field site model. Both modelling approaches indicated size of stream system to be an important influence on occurrence of juvenile grayling in the Williston watershed. River length required for suitable river habitat for salmonids has not previously been identified, but should be factored into future management plans when evaluating the impact of proposed hydroelectric dams and subsequent flooding of river systems.     

The potential exceptional role of a small Baltic boulder reef as a solitary habitat in a sea of mud

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The isoelectric point, a key to understanding a variety of biochemical problems: a minireview

We address the importance of the isoelectric point (IEP) of proteins and membrane components such as phospholipids for our understanding and interpretation of isoforms and opposite charge interactions in the formation of complexes. Five examples drawn from the literature are newly approached from the IEP point of view to clarify general principles.     

Acoustic analysis of volume variation in a bag-net within a set-net

An experiment designed to measure the volume variation of a bag-net within a set-net was conducted in Jaran Bay, Kosung, Korea. Three radio-acoustic-linked positioning (RAP) buoys, a time controller with a personal computer and seven pingers were used to measure the volumes of the bags. During the April neap tide, the minimum and maximum volumes of the bag-net were 4173 m³ (at 17.00 h) and 4757 m³ (12.00 h), respectively. The average current directions and speeds were 99.9°, 12.9 cm/s and 104.0°, 2.4 cm/s, respectively. During the spring tide, the minimum and maximum volumes of the bag-net were 2016 m³ (18.30 h) and 4454 m³ (15.00 h), respectively. The average current directions and speeds were 315.6°, 16.1 cm/s and 289.0°, 5.7 cm/s, respectively. The minimum (2016 m³) and maximum (5568 m³) volumes of the bag-net were observed during the period when the spring tide changed to the neap tide.     

Evaluation of a portable electrode array for a resistivity fish counter

Abstract This study aimed to develop and test an electrode array for a resistivity fish counter that could be easily installed in a small river without a weir. An electrode array consisting of three steel cables laid in parallel across the stream channel and connected to a microprocessor-based counter was tested in an Atlantic salmon, Salmo salar L., spawning tributary. The accuracy of the counter was assessed by observing fish movements with closed-circuit television. Most salmon moving upstream were registered correctly (90% overall). Detection of downstream movement was less reliable (60% overall), as a consequence of downstream swimming behaviour. The accuracy of the downstream count was improved by tensioning the cable electrodes, but remained lower than that of the upstream count. Since salmon swam repeatedly up- and downstream, this discrepancy resulted in an overestimate of the net upstream count. The accuracy of the downstream count needs to be improved before a bed-mounted electrode array could be used for routine salmon counting.