僧帽牡蛎碱性磷酸酶性质的研究

从僧帽牡蛎分离纯化出一种碱性磷酸酶,采用Sephadex G-200凝胶过滤柱层析法测得该酶分子量为1.57×10~5.以对硝基苯磷酸为底物,该酶的最适温度为43℃,最适pH值为10.0.在37℃和pH值为10.0的条件下,酶催化反应的米氏常数(K_m)为9.68×10~(-4)mol/dm~3,活化能(E_a)为45.85kJ/mol,温度系数(Q_(10))为1.80(30～40℃).对该酶的热稳定性研究表明:其在45℃以下较为稳定,50℃以上明显失活,并测定了该酶的热失活速度常数.金属离子对酶活力的效应研究结果表明:Mg~(2+)、Mn~(2+)、Ca~(2+)、Co~(2+)对酶有激活作用,Ag~+、Hg~(2+)、Zn~(2+)表现为抑制作用,Cd~(2+)几乎没有影响.Mg~(2+)是较有效的激活剂,表现为部分非竞争性效应,激活常数(K_a)为2.48×10~(-4)mol/dm~3,有Mg~(2+)存在时该酶活性中心的转换数量是无Mg~(2+)时的4.0倍.几种效应物对酶的抑制机理研究表明:HPO_4~(2-)、HAsO_4~(2-)、Cys为竞争性,其抑制常数分别为1.61、1.18和0.15mmol/dm~3;L-Phe为反竞争性,抑制常数为3.26mmol/dm~3.     

海洋芽孢杆菌Bacillus sp. CAMT22370所产葡萄糖氧化酶的分离纯化与表征

前期筛选获得一株海洋源Bacillussp.CAMT22370高产葡萄糖氧化酶菌株,本研究进一步通过离子交换及分子筛层析使Bacillus sp. CAMT22370葡萄糖氧化酶纯化了2.12倍,酶活得率32.37%,比活力达到8.47 U/mg,分子质量约为50 kDa。酶学特性研究表明,此酶在低温下有较好的催化活性,最适温度为30℃,超过30℃时酶活迅速下降, 45℃保温2 h活力降至75%左右;最适pH为6.0,在5.0以下和7.0以上时酶活迅速降低, 9.0时损失35%; Ca~(2+)和Mn~(2+)有明显激活作用, Na~+和Zn~(2+)有轻微激活作用, K~+和Mg~(2+)对酶活无显著影响,而Fe~(2+)、Fe~(3+)和Cu~(2+)对酶活有明显抑制作用,酶动力学参数Vmax及Km值分别为26.85 (μmol/(L·min))和109.26 (μmol/(L·min)),以上酶学特性与现有酶制剂存在明显不同,研究结果为该葡萄糖氧化酶的开发利用奠定了初步基础。     

深海细菌Pseudomonas sp. IOFA1中甲醛歧化酶FM11的纯化及性质研究

从印度洋深海沉积物中分离出一株能够高效降解甲醛的假单胞菌(Pseudomonas sp.IOFA1),该菌株可在40 min内完全降解100 mg/cm~3的甲醛.通过阴离子交换柱层析对菌株破碎液中的粗酶液进行纯化,得到一条具有甲醛降解活性的单一蛋白条带.经MALDI-TOF/TOF质谱鉴定,该蛋白为甲醛歧化酶蛋白(FM11).序列分析表明FM11与其它物种已知醛脱氢酶的氨基酸序列最高相似性为65%;结构域分析表明FM11含有2个保守的锌结合位点和一个NAD结合位点.酶学性质分析结果显示,FM11的最适反应温度为40℃,在25~50℃范围内能够保留95%以上的酶活力,尤其是在高于70℃的高温环境下仍保有约50%的活力,表现出广泛的温度适应性.FM11具有非常广泛的最适反应pH值范围(pH值为5~9),且在pH值4和pH值10时仍分别保留50%和80%左右的酶活力,表现出广泛的pH适应性.在pH值为5~7的条件下孵育1 h后,FM11仍保有97%以上的酶活,且在pH值为7~10条件下孵育1 h后,仍保留90%以上的酶活,表现出极好的酸性和碱性稳定性.Ca~(2+)、Mg~(2+)在低浓度时(0.5 mmol/dm~3)对FM11有较明显的激活作用.这些研究为我们进一步开发Pseudomonas sp.IOFA1甲醛歧化酶在降解甲醛方面的用途奠定了基础.     

海洋交替单胞菌（Alteromonas sp. Y-389）普鲁兰酶基因的异源表达与酶学性质分析

以海洋交替单胞菌(Alteromonas sp. Y-389)的基因组为模板,设计普鲁兰酶基因的引物并进行PCR扩增。将目的基因连接到载体pET-28a(+)后进行测序并进行生物信息学分析。结果显示,基因的开放阅读框全长4 347 bp,编码1 449个氨基酸的普鲁兰酶,为Ⅰ型普鲁兰酶,有4个保守的结构域,属于糖苷水解酶家族13;将重组质粒转化到大肠杆菌BL21(DE3),成功诱导出了可溶性的目标蛋白;随后对其酶学性质进行测定与分析,结果表明,该酶的比活力为54.53 U/mg,其最适的反应温度为35℃,最适pH为6.0,Na~+和Mn~(2+)对该酶的活性具有明显的促进作用,而Cu~(2+),Zn~(2+)与Fe~(2+)对其活性的抑制作用比较明显;其酶学参数K_m和V_(max)分别为3.12 mg/mL,228.8 U/mg,将为今后的规模化生产提供数据支持。     

产甲壳素酶菌株的发酵条件、酶的分离纯化及酶学性质研究

利用甲壳素为唯一碳源从土壤中筛选得到1株产甲壳素酶活力较高的菌株HD002,初步鉴定其为Massilia属。确定菌株产甲壳素酶的最适培养基组成为(g/L):(NH4)2SO45,K2HPO40.7,KH2PO40.3,MgSO4.7H2O 1,胶体甲壳素10;最适产酶培养条件为:培养基起始pH值6.0,培养时间192 h,发酵液酶活力达1.314 U/mL。采用70%饱和度硫酸铵盐析、DEAE-Sepharose F.F阴离子交换层析、Sephadex G-75凝胶过滤层析对该甲壳素酶进行纯化,SDS-PAGE证明达到电泳纯。该甲壳素酶的分子量为60.2 kDa,最适反应温度为55℃,最适反应pH值为5.0,Cu2+和Fe3+对酶活力有明显的抑制作用,Ca2+和Na+对酶活力有明显的促进作用。     

中国鲎内脏N-乙酰-β-D-氨基葡萄糖苷酶的分离纯化及其酶学性质

以中国鲎(Tachypleus tridentatus)内脏为材料,分别通过30%和80%饱和度(NH_4)_2SO_4沉淀分级分离、葡聚糖凝胶柱Sephadex G-200层析、以及离子交换柱DEAE-32层析,从而获得N-乙酰-β-D-氨基葡萄糖苷酶(EC3.2.1.52,NAGase),经PAGE检定达到电泳纯,酶的比活力为505.21 U/mg。SDS-PAGE显示一谱带,测得该酶蛋白亚基分子量为121.5kDa,等电聚焦电泳法测得酶的等电点pI为6.01。以pNP-β-D-GlcNAc为底物,研究NAGase催化反应的动力学参数。结果表明:中国鲎NAGase的最适pH、最适温度、K_m和V_(max)分别为5.4和55℃、0.421 mmol/L和13.158μmol/L·min~(-1);酶在pH=4.5~7.0范围内较稳定,在20~50℃之间具有较好的热稳定性。酶在276nm波长处有紫外吸收峰,在232.2nm波长的内源荧光激发下,酶内源荧光发射光谱峰在330.9nm波长处。Na~+,K~+和Li~+对NAGase活力没有影响,Ca~(2+)、Ba~(2+)和Co~(2+)对NAGase有不同程度的激活作用,Co~(2+)对NAGase的激活作用较大,5mmol/L Co~(2+)能使NAGase活力提高41.67%;Mg~(2+)、Cu~(2+)、Zn~(2+)、Mn~(2+)、Fe~(3+)、Al~(3+)、Pb~(2+)、Ag~+、Cd~(2+)和Hg~(2+)等10种金属离子对NAGase活力有不同程度的抑制作用,5mmol/L Cd~(2+)可使酶活力丧失85.60%,而Hg~(2+)对酶的抑制作用最强,0.1mmol/L Hg~(2+)可使酶活力丧失90.10%。EDTA对酶活力没有影响,推断中国鲎NAGase属于非金属酶类。     

一株源于红树林根际土壤产纤维素酶芽孢杆菌的鉴定及其酶学性质分析

从红树林根际土壤中筛选出一株产纤维素酶的革兰氏阳性菌,通过形态、生化特征及16S r RNA基因序列分析鉴定为枯草芽孢杆菌(Bacillus subtilis)。对其产纤维素酶能力进行分析,发现该菌株所产纤维素酶的最适反应温度为55°C,最适p H为6。酶学性质分析发现,在反应温度为55°C时,该菌株产生的纤维素酶稳定性最好,保温120min后仍具有80%以上的相对酶活力;在p H为5—7时,酶活性相对稳定,相对酶活力均维持在80%左右。Cu~(2+)对该酶有一定的抑制作用,Zn~(2+)、Fe~(3+)、Mg~(2+)、Ca~(2+)、Fe~(2+)、Mn~(2+)和Co~(2+)均对酶有一定的促进作用。本研究显示这株芽孢杆菌具有良好的应用前景,为该菌株在水产饲料添加剂中的进一步研究与应用提供参考依据。     

海藻工具酶研究Ⅱ.褐藻酸酶的制备、性质及对裙带菜细胞的解离研究

褐藻酸降解菌埃氏交替单胞菌(Alteromonas espejiana)菌株Al01,通过发酵培养制备褐藻酸酶.该酶作用的最适pH为7.0,最适温度为40℃,最适底物浓度为1%～2%;在离子浓度为0.5mmol/dm3时,Mn2+对酶促反应稍有促进作用,Ca2+、Hg2+则有明显的抑制作用.该酶用于裙带菜单细胞和原生质体解离时,以酶液组成为褐藻酸酶1%、纤维素酶1%,45×10-3的NaC1为渗透剂,酶解温度25℃,pH7.0,酶解3-4h效果最好.     

印尼热泉菌产热稳定褐藻胶裂解酶的酶学性质研究

以印尼热泉菌为材料,采用以褐藻酸钠为唯一碳源的培养基筛选产褐藻胶裂解酶的菌株,通过16S rDNA序列对产酶菌株进行种属鉴定,并通过硫酸铵沉淀、离子交换层析和凝胶过滤层析从高产酶菌株Aly-B5发酵液上清中纯化获得褐藻胶裂解酶,并对该酶的酶学性质进行了研究。结果表明:高产酶菌株经鉴定为Bacillus属;聚丙烯酰胺凝胶电泳(Sodium dodecyl sulfate polyacrylamide gel electrophoresis,SDS-PAGE)显示纯化的褐藻胶裂解酶Aly-B5分子量为45kDa,该酶最适作用温度为65°C,75°C时的半衰期为110min,最适pH为7.0;Zn~(2+)、Mg~(2+)、Fe~(2+)对酶活具有明显的抑制作用,但该酶对乙二胺四乙酸(ethylene diamine tetraacetic acid,EDTA)并不敏感;同时,底物特异性实验表明Aly-B5是一种偏好降解聚古罗糖醛酸(poly G)的双功能裂解酶,降解产物主要是二糖。该酶优良的耐热性使其在海藻高值化利用领域具有潜在的应用前景。     

海洋细菌QY202产κ-卡拉胶酶的分离纯化和性质研究

为获得高效降解卡拉胶菌株,从青岛太平角海域采集的角叉菜表面分离到1株高产κ-卡拉胶酶的海洋交替假单胞菌(Pseudoalteromonas sp.)QY202,经硫酸铵沉淀、脱盐、DEAE阴离子交换层析等步骤从该菌株发酵液上清中分离纯化得到1种专一性降解κ-卡拉胶的κ-卡拉胶酶,并研究了该酶的基本酶学性质.结果表明该酶被纯化了23.1倍,回收率为43.9%,分子量大小为33.2 kDa.酶的最适反应温度为40 ℃,最适反应pH为8.0,在0～40 ℃,pH=7.0～8.0之间酶活力较稳定.酶对底物κ-卡拉胶的米氏常数Km值为1.6 mg/mL.Na~+、K~+对酶活有促进作用,而Hg~(2+)、Cu~(2+)强烈抑制酶的活性.酶解κ-卡拉胶的主产物为硫酸新κ-卡拉二糖和硫酸新κ-卡拉四糖.     

Microbial Biomass and Organic Nutrients in the Deep-Sea Sediments of the Central Indian Ocean Basin

In order to assess the impact of deep-sea mining on the in situ benthic life, we measured the microbial standing stock and concentration of organic nutrients in the deep-sea sediments of the Central Indian Ocean Basin in the Indian pioneer area. Sediments were collected using box core and grab samples during September 1996. The total bacterial numbers ranged from 10 10 -10 11 cells per g -1 dry weight sediment. There was a marginal decrease in the number of bacteria from surface to 30 cm depth, though the subsurface section registered a higher number than did the surface. The highest numbers were encountered at depths of 4-8 cm. The retrievable number of bacteria were two orders less in comparison with the direct total counts of bacteria. An almost homogeneous distribution of bacteria, total organic carbon, living biomass, and lipids throughout the depth of cores indicates active microbial and benthic processes in the deep sea sediments. On the other hand, a uniform distribution of total counts of bacteria, carbohydrates, and total organic carbon in all the cores indicates their stable nature and suggests that they can serve as useful parameters for long-term monitoring of the area after the benthic disturbance. Further studies on temporal variability in this region would not only verify the observed norms of distribution of these variables but would also help to understand restabilization processes after the simulated benthic disturbance.     

海上大直径钢管桩打桩过程中桩周土强度弱化研究

海上大直径钢管桩打桩过程中,桩周土体受到强烈扰动而发生强度弱化,掌握桩周土体强度弱化规律对于准确预测打桩过程、保证工程安全具有重要意义。为研究土体强度弱化规律,开展了环剪试验模拟打桩对桩周土体的扰动,测试土体强度随剪切速率的变化规律,建立了描述土体强度弱化规律的拟合公式,引入到打桩分析软件中。研究结果表明：土体的强度折减程度不仅与土体本身的性质有关还受到土体的埋深和剪切速率的影响,埋深越深土体强度折减程度越低,剪切速率越高土体强度折减越高,在打桩分析中可采用这里推荐的线性折减方法来模拟不同深度处土体强度的折减规律。     

Geoacoustic Inversion via Genetic Algorithm and Its Application to Manganese Sediment Identification

An acoustic inversion method using a wide-band signal and two near field receivers is proposed and applied to multiple layered seabed models including a manganese sediment. The inversion problem can be formulated into a probabilistic model comprised of signals, a forward model, and additive noise. The forward model simulates wide-band signals, such as chirp signals, and is chosen to be the source-waveletconvolution plane wave modeling method. The wavelet matching technique, using weighted least-squares fitting, estimates the sediment sound-speed and thickness on which determination of the possible numerical ranges for a priori uniform distribution is based. The genetic algorithm is applied to a global optimization problem to find a maximum a posteriori solution for determined a priori search space. Here the object function is defined by an L 2 norm of the difference between measured and modeled signals. Not only the marginal pdf but also its statistics are calculated by numerical evaluation of integrals using the samples selected during importance sampling process of the genetic algorithm.     

Phylogenetic diversity and phenotypic characterization of cultivable bacterioplankton isolated from polar oceans

A set of 27 marine planktonic bacteria isolated from the polar regions was characterized by 16S rDNA sequencing and physiological and biochemical testing. More than half of these bacteria were positive for caseinase, gelatinase and 13-glucosidase, and could utilize glucose, maltose or malic acid as carbon source for cell growth. Twelve isolates expressed nitrate reduction activities. Except for one antarctic isolate BSwlO175 belonging to Actinobacteria phylum, these isolates were classified as γ-Proteobacteria, suggesting that γ-Proteobacteria dominated in cultivable marine bacterioplankton at both poles. Genus Pseudoalteromonas was the predominant group in the Chukchi Sea and the Bering Sea, and genus ShewaneUa dominated in cultivable bacterioplankton in the Prydz Bay. With sequence similarities above 97%, genus Psychrobacter was found at both poles. These 27 isolates were psychrotolerant, and significant 16S rDNA sequence similarities were found not only between arctic and antarctic marine bacteria （ 〉 99% ）, but also between polar marine bacteria and bacteria from other aquatic environments （ ≥ 98.8% ）, including temperate ocean, deep sea, pond and lake, suggesting that in the polar oceans less temperature-sensitive bacteria may be cosmopolitan and have a bipolar, even global, distribution at the species level.     

Deep-Sea Sediment Compression Curves: Some Controlling Factors, Spurious Overconsolidation, Predictions, and Geophysical Reproduction

Oedometer tests have been carried out on 70 undisturbed surficial clays (at approximately 250 mm below the mudline), mostly collected by free-fall corers from sites widely scattered throughout the deep-sea North Atlantic. Acoustic measurements were also made, initially on contiguous samples and ultimately on the same sample using a geophysically instrumented oedometer which also collected electrical resistivity data. Apart from those quiescent areas below the carbonate compensation depth, such as north of the West Indies where very fine clays exist, most of the samples are silty clays whose geotechnical-geophysical properties are dependent on the type of clay minerals present (and their ability to take in moisture), the sand-size fraction, and the quantity of carbonate present. Thus the pure clays have high compressibilities which decrease on the addition of coarse particles, while the converse is true for the acoustic parameters, these increasing with the sand fraction. Using the notion of the intrinsic compression line for all samples, and comparison to it of the measured compression curves, it is clear that, contrary to some previously held ideas, most deep-sea clays are normally consolidated; the addition of carbonate has the effect of creating an open, stronger sediment skeleton. Interestingly, where information is available, the variation with depth of a sample's acoustic velocity follows the void ratio pressure relationship of the compression curve. This allows the construction of an in-situ sediment compression curve using the in-situ geophysical observations.     

INFORMATION FOR AUTHORS

<正>Acta Oceanologica Sinica(AOS)is a comprehensive academic journal edited by the Editorial Committee of Acta Oceanologica Sinica and is designed to provide a forum for important research papers of the marine scientific community which reflect the information on a worldwide basis.The journal publishes scholarly papers on marine science and technology,including physics,chemistry,biology,     

Evaluation of the Risk of Marine Slope Instability: A Pseudo-3D Approach for Application to Large Areas

This article presents a methodology developed to evaluate the instability of submarine slopes that extend over a large area. Special attention was paid to (1) the complex geometry (bathymetry) and the expanse of the slope, (2) the heterogeneity of the sediment, and (3) the distribution of the pore pressure. The safety factor was considered as a spatially varying quantity. The General Formulation (GLE, Fredlund and Krahn 1977), which fully satisfies equilibrium conditions, was used for evaluating the stability of the marine slope. The submarine slope failure, which occurred on 16 October 1979 during the construction of the new Nice airport, was studied in order to test the developed model. Geotechnical parameters were taken from experimental tests carried out by IFREMER on 19 cores extracted at different depths (from 27 m to 1300 m) (Cochonat, Bourillet, and Savoye, 1993; Mulder et al., 1994). Many scenarios were proposed in order to explain the cause of the Nice slope failure (Habib, 1994). In this article, two of those scenarios were tested. Simulation results are presented and discussed.