Adsorption and desorption of proteins and polyamino acids by clay minerals and marine sediments

The adsorption and desorption of proteins and polyamino acids on illite, montmorillonite, goethite, and marine sediments was investigated. Three ¹⁴C-labeled hydrophilic proteins, Rubisco from C. reinhardtii, and GroEL and GroES from genetically modified Escherichia coli, were synthesized and purified for this study. The proteins were strongly and rapidly adsorbed by the clay minerals and marine sediments, and much of the adsorbed protein was not readily desorbed. Sodium dodecyl sulfate (SDS) extraction and separation by SDS-polyacrylamide gel electrophoresis (PAGE) and sucrose density gradients showed that Rubisco and GroEL were adsorbed on illite and sediments in their original forms. The apparent adsorption partition coefficients of the proteins were on the order of 10² l/kg on illite, 10³–10⁴ l/kg on goethite and montmorillonite, and 200 and 75 l/kg on Skan Bay and Resurrection Bay (Alaska) sediments, respectively. These partition coefficients are sufficiently large to permit sedimentary protein preservation via an adsorptive mechanism. Generally, basic polyamino acids had greater adsorption partition coefficients than acidic polyamino acids. Molecular size did not affect the electrostatic interaction between polyamino acids and mineral surfaces. Adsorption of bovine serum albumin (BSA) and negatively charged polyamino acids inhibited Rubisco adsorption, while positively charged cytochrome c and polyamino acids increased Rubisco adsorption. These results indicate that electrostatic interactions dominated in protein adsorption.     

光强和营养盐对伪矮海链藻昼夜节律变化的影响

于1989年1月—1989年8月,用连续培养和半连续培养方法进行了伪矮海链藻碳水化合物、蛋白质和生化组成比率的日变化同光强、营养盐关系的研究。结果表明,碳水化合物、碳水化合物同叶绿素a和蛋白质的比值均呈现光照期升高而黑暗期降低的昼夜节律;蛋白质在光照期和黑暗期均有合成,但是,光照期的增长速率大于黑暗期的;蛋白质同叶绿素a的比值无明显的日变化。各指标的日变化幅度均随光强的增强和营养盐供应速率的增加而升高,因此,光和营养盐是影响各指标日变化的重要因素。     

化学融雪剂对小球藻生长和生理特性的影响

寒冷地区道路表面除雪化冰使用的化学融雪剂随着地表径流进入水体,将影响水生态系统中的水生生物正常生长,并破坏水生态系统平衡。为研究化学融雪剂对水生生物的毒性效应,分析了不同浓度有机融雪剂对小球藻生长特征、藻细胞光和色素、蛋白质及多糖含量的影响。结果表明：浓度为2 g·L^-1时有机融雪剂对小球藻生长无明显影响,当融雪剂浓度为4 g·L^-1时,小球藻细胞生长表现出明显抑制效应,且抑制效应随融雪剂浓度增加呈显著上升趋势(P<0.01);当融雪剂浓度小于4 g·L^-1时,有机融雪剂对小球藻细胞内叶绿素a合成有明显促进作用,但随着有机融雪剂处理浓度升高,藻细胞内叶绿素a含量逐渐下降。当融雪剂的处理浓度大于4 g·L^-1的时候,藻细胞内蛋白质和多糖含量与对照组相比呈显著下降趋势(P<0.01)。这说明融雪剂浓度高于4 g·L^-1时会抑制水体中小球藻正常生长繁殖,破坏藻体细胞,最终导致水生态系统平衡被破坏。     

Polybrominated diphenyl ethers do not affect metamorphosis but alter the proteome of the invasive slipper limpet Crepidula onyx

Man-made polybrominated diphenyl ethers (PBDEs) used as flame retardants in various consumer products may be harmful to marine organisms. Larvae of some marine invertebrates, especially invasive species, can develop resistance to PBDEs through altered protein expression patterns or proteome plasticity. This is the first report of a proteomics approach to study BDE-47 induced molecular changes in the invasive limpet Crepidula onyx. Larvae of C. onyx were cultured for 5 days (hatching to metamorphosis) in the presence of BDE-47 (1 μg L⁻¹). Using a 2-DE proteomics approach with triple quadrupole and high-resolution TOF-MS, we showed that BDE-47 altered the proteome structure but not the growth or metamorphosis of C. onyx larvae. We found eight significant differentially expressed proteins in response to BDE-47, deemed the protein expression signature, consisting of cytoskeletal, stress tolerance, metabolism and energy production related proteins. Our data suggest C. onyx larvae have adequate proteome plasticity to tolerate BDE-47 toxicity.     

海洋溴酚衍生物LXQ-5的PTP1B的结合作用和体内降糖活性研究

二型糖尿病是一种代谢性疾病,对人类健康造成了极大的威胁。蛋白酪氨酸磷酸酶1B(PTP1B)是胰岛素信号通路中的负调控蛋白酶,是二型糖尿病治疗的重要研究靶点。本文利用酶动力学实验和分子互作技术检测了溴酚衍生物LXQ-5的PTP1B抑制作用类型和体外特异性结合作用的特点。测定糖尿病小鼠口服给药后的空腹血糖水平和血清中糖化血红蛋白以及糖化血清蛋白水平等糖尿病相关指标的变化,研究了化合物LXQ-5在小鼠体内的降糖活性。实验结果表明, LXQ-5是PTP1B的一种非竞争性抑制剂,且在体外能够与PTP1B特异性结合。LXQ-5在糖尿病小鼠体内能显著的降低空腹血糖水平和血清中糖化血红蛋白、糖化血清蛋白含量,在新型降糖药物的研发方面具有重要的研究价值。     

Application of pyrolysis–GC/MS for the characterisation of suspended particulate organic matter in the Mediterranean Sea: a comparison with the Black Sea

Suspended particulate organic matter (SPOM) both from the oligotrophic Mediterranean and from the more productive Black Sea has been analysed by pyrolysis–gas chromatography/mass spectrometry (Py–GC/MS) and by such conventional techniques as elemental analysis and fluorometry. The data achieved by the two approaches generally confirmed and complemented each other. The relative concentrations of pyrolysis products (termed markers) characteristic of chlorophyll (CHL), lipid, carbohydrate (CBH), and protein (PROT) components of SPOM have been determined. The vertical distribution of lipid markers was more uniform in the Mediterranean surface waters. Their relative abundance increased rapidly in the oxycline of the Black Sea, reaching their highest levels in the suboxic zone, where the protein composition of the SPOM changed significantly. In both seas, the relative concentrations of CHL markers increased consistently within the CHL maximum zone where the CBH markers were less abundant. No lignin markers from terrestrial vegetation were detected at significant levels in the pyrograms.     

动物蛋白酶解物与无机氮、磷源的组合对小球藻(Chlorella vulgaris)生长和生化指标的影响

采用三因素不完全正交设计方法,通过对细胞密度、干重、叶绿素和可溶性蛋白质含量的测定.确定了N、P源与蛋白酶解物之间的最佳组合,研究碳酸氢铵作为N源、磷酸二氢钠作为P源与动物蛋白酶解物之间的组合对小球藻生长和生化指标的影响.结果表明:(1)动物蛋白酶解物与N、P源间存在互作效应--影响小球藻细胞密度、叶绿素的主要因素分别是P浓度、蛋白酶解物浓度;而N浓度及蛋白酶解物浓度同时对小球藻的蛋白质含量起主要作用;(2)同时添加N 50mg/L、P 4mg/L、蛋白酶解物12ml/L能显著提高小球藻生物量,促进叶绿素和藻体蛋白质的合成(P<0.05).     

变蛋中微量铅的催化示波极谱法测定

用催化示波极谱法测定变蛋中微量铅 ,提出在HCl-酒石酸 -KI -抗坏血酸体系中 ,用催化示波极谱法测定变蛋中微量铅。在此体系中 ,Pb(ⅠⅠ )于 -0 54V处产生一个非常灵敏的吸附催化波。铅含量在 0 0 2 5～ 3 μg·mL-1 范围内波高与浓度呈良好线型关系 ,相关系数r为 0 9976,检测限为0 0 2 5μg·mL-1 。样品经HNO3 -HClO4消化和分解 ,蒸至近干后 ,再用HCl溶解 ,在HCl-酒石酸 -KI体系中测定 ,回收率为 ( 98± 2 ) %。此方法具有灵敏、准确、快速等特点 ,适用于变蛋及其他食品、人发等样品中微量铅的测定     

钝顶节旋藻(Arthrospira platensis)2-DE分析蛋白制备方法改进及试用于螺旋手性差异蛋白研究

采用冻融破壁和蛋白质分步提取等技术,对钝顶节旋藻2-DE分析蛋白制备方法作了改进,并试用于螺旋手性差异表达蛋白研究,结果表明:(1)钝顶节旋藻细胞反复冻融5次完全破碎,用Tris-HC1提取液提取3次可提得近87％的水溶性蛋白,再用SDS(十二烷基磺酸钠)提取液可从沉淀中提得水不溶性蛋白;(2)将上述水溶性和水不溶性蛋白分别用TCA/丙酮法纯化后作双向电泳(2-DE)分析,可辨蛋白点分别达500和760多个,而两者间的匹配率仅7％,说明分离度好且质量高;(3)利用此法获得了13个与钝顶节旋藻螺旋手性相关的候选蛋白,其中5个为首次发现,候选蛋白功能涉及光合作用、能量代谢、物质外排和环境适应等;(4)本文所建的“冻融破壁分步提纯法”与现有节旋藻2-DE蛋白制备方法相比,具有操作简便、仪器设备简单、成本低廉及信息量更丰富等优点.     

Photosynthate allocation in a temperate sea over an annual cycle: the relationship between protein synthesis and phytoplankton physiological state

The seasonal and vertical variations in the patterns of photosynthate allocation into biomolecules by natural phytoplankton assemblages were determined, together with their species composition, in a coastal station of the central Cantabrian Sea (southern Bay of Biscay). Chlorophyll-a concentration ranged from values below 20 mg m⁻² in winter to values above 80 mg m⁻² during spring and during an upwelling event in summer. Low primary production rates (<300 mgC m⁻² d^-1) were measured during winter and during summer stratification periods. The rate of C fixation during summer upwelling conditions exceeded 3500 mgC m⁻² d⁻¹. In terms of photosynthate partitioning, proteins were the dominant fraction, as they typically accounted for >30% of total photo-assimilated C, with polysaccharides and low molecular weight metabolites showing incorporation percentages around 10–30%. Relative C incorporation into lipids was generally <15%. Recurrent patterns of vertical variability in photosynthate partitioning were observed: the relative synthesis of proteins increased toward the bottom of the euphotic zone, whereas the relative C incorporation into polysaccharides and lipids tended to be higher near the surface. When primary production decreased, the synthesis of proteins was maintained more than that of other molecules. Throughout the year, the relative synthesis of proteins was inversely correlated with phytoplankton biomass, production and growth rate. The conservation of protein synthesis under growth-limiting conditions and the enhancement of lipid and polysaccharide synthesis when irradiance is high seem to constitute general patterns of photosynthate partitioning in marine phytoplankton. In our study, these patterns represented metabolic strategies of phytoplankton in response to changing environmental factors, rather than the effect of variations in the species composition of the community.