针对点突变癌基因T24－ras转录物的核酶性质研究

通过改变核酶体外反应的各种条件,对针对点突变癌基因Ｔ２４－ｒａｓ转录物的核酶Ｒ８的性质进行了系统研究,结果表明：Ｒ８切割底物反应的最适ｐＨ值约为８．２,最适温度为５０℃,反应必须有镁离子参加,生理环境中常见的阴离子对反应没有影响,一些变性剂能够促进反应的进行,Ｒ８切割底物的反应为二级反应。     

针对点突变癌基因T24—ras转录物的核酶性质研究

通过改变核酶体外反应的各种条件,对针对点突变癌基因Ｔ２４－ｒａｓ转录物的核酶Ｒ８的性质进行了系统研究,结果表明：Ｒ８切割底物反应的最适ｐＨ值约为８．２,最适温度为５０℃,反应必须有镁离子参加,生理环境中常见的阴离子对反应没有影响,一些变性剂能够促进反应的进行,Ｒ８切割底物的反应为二级反应。     

生物制造“细胞工厂”的设计与组装

以化石资源为原料的化学品制造行业在消耗不可再生资源的同时,还对生态环境造成了破坏,这给以可再生资源为原料的生物制造带来了发展机遇。与传统化工制造不同,生物制造把细胞作为"生产车间","车间"内每一道工序由酶催化完成。"细胞工厂"除了反应条件温和,还具有较强的可塑性,可根据需求调整或者重构代谢途径来合成各种目标化学品。"细胞工厂"的设计过程遵循如下的准则:1)构建一条由原料到产品的最优合成途径;2)平衡代谢途径中每步反应的代谢流,使该途径代谢通量远高于细胞基础代谢;3)足量地供应合成途径的前体,多个前体根据需要调整供应比例;4)酶促反应往往有各种辅因子的参与,顺畅的代谢通路需要平衡或者再生各种辅因子;5)通过遗传改造或者工艺改进解除产物和代谢中间体的反馈抑制,以获取更高的产量。     

基于万古霉素建立荧光酶联免疫吸附法检测金黄色葡萄球菌

以猪IgG作为捕获抗体固定金黄色葡萄球菌,修饰有万古霉素的量子点荧光微球作为"检测抗体",建立荧光酶联免疫吸附法检测金黄色葡萄球菌。文中制备了平均粒径为100 nm的量子点荧光微球并与万古霉素偶联;摸索了反应最佳盐离子浓度为0.01 mol/L,反应最佳pH为6.0。在该实验条件下,金黄色葡萄球菌的检测灵敏度为104 CFU/m L,与其他致病菌无交叉反应。以上结果表明,该方法可用于快速检测金黄色葡萄球菌,为金黄色葡萄球菌的临床监控和食品检测提供参考。     

毒死蜱和阿维菌素对塔六点蓟马功能反应的影响

室内采用叶碟饲养的方法,以山楂叶螨Tetranychus viennensis Zacher卵为猎物研究了苹果园常用药剂毒死蜱和阿维菌素对塔六点蓟马Scolothrips takahashii Prisener功能反应的影响。结果表明：毒死蜱和阿维菌素对塔六点蓟马的功能反应均有明显的影响。在推荐剂量下阿维菌素可显著降低雌雄两性蓟马的瞬时攻击率,处置时间则分别比对照增加213.36%和19.74%,且雌性的功能反应由HollingⅡ型改变为HollingⅢ型;在半推荐剂量下两性蓟马的瞬时攻击率与对照差异不明显,而雌性蓟马的处置时间则比对照增加133.72%。毒死蜱处理则不影响塔六点蓟马的功能反应类型,在推荐剂量和半推荐剂量下均可使雌雄两性蓟马的瞬时攻击率显著降低,雌性蓟马的处置时间比对照分别增加85.62％和71.97%,而雄性蓟马在两个剂量下则分别增加55.92％和38.20%,与对照的差异均达显著水平。     

类囊体膜的能化状态与光合磷酸化反应活化能的关系

在0～28℃范围内测定了菠菜离体叶绿体的光合电子传递和磷酸化反应的活化能。发现叶绿体处于解联状态时,Hill反应的Arrhenius图在7～8℃处呈现出活化能改变的折点。叶绿体在光下的能化状态由于加入ADP及Pi进行磷酸化反应而消除时,活化能的折点依然在8℃附近呈现,表明活化能折点的出现与光合膜能化状态消失有关。不同光系统的电子传递和磷酸化反应的Arrhenius图表明,活化能折点的出现主要发生在与PS-Ⅱ有关的反应中,而与PS-Ⅰ反应的关系不大。     

关于茚三酮法测定脯氨酸含量中脯氨酸与茚三酮反应之探讨

测定脯氨酸含量的方法通常用磺基水杨酸或酒精提取再与酸性茚三酮反应生成稳定的红色产物,然后于波长520 nm处比色,通过标准曲线或回归方程计算脯氨酸的含量.关于其测定原理,一些植物生理学实验指导明确指出:"在酸性条件下,脯氨酸和茚三酮反应生成稳定的红色产物,用比色法于520 nm波长下测定脯氨酸的含量."[1,2]然而,在各种生物化学教科书[3～6]中描述氨基酸茚三酮反应时,则为:"在弱酸性条件下,2个亚氨基酸--脯氨酸和羟脯氨酸与茚三酮反应并不释放NH3,而直接生成黄色化合物,其结构式如图1,最大光吸收在440 nm."为了对两种教材中不同的说法进行比较,我们查阅了各种文献,设计一些试验并进行了验证.     

生物无机化学的新动向

介绍了近几年生物无机化学发展的新动向,特别是各种与DNA,RNA相结合的金属调节蛋白,在核酸基因表达及调控中的作用。金属蛋白与金属酶的研究,由于基因工程、金属络合物探针和肽链的人工合成等各种新思想、新方法的引入,并且把各种金属蛋白反应作为生物体中的一类反应进行研究(例如:生物体的电子传递反应,金属离子在生物分子间的转移反应,小分子加合反应等等),使研究进入一个新的台阶。     

细胞周期检验点与肿瘤发生之间关系的研究进展

DNA损伤反应引起的基因组不稳定性并不足以导致肿瘤发生,还需要一些协同突变促进肿瘤的生长或存活,因此,基因组结构不稳定和周期检验点突变失活是肿瘤发生的重要因素。与正常细胞不同,肿瘤细胞中细胞周期检验点反应缺陷,当肿瘤细胞遭受基因毒药物损伤时,可通过激活周期检验点反应阻滞细胞周期进程,加强损伤修复,导致耐药表型的产生。因此,寻找特异性的检验点抑制剂来加强化疗药物或辐射对肿瘤细胞的杀伤效应,已成为肿瘤治疗的一个研究方向。     

橡胶树死皮病的发生机理和假说

在阐述橡胶树死皮病的概念和已有关于死皮病因及其机理假说基础上,根据分子生物学新进展提出了死皮病发生的新现点.死皮病是可由生物、生理多种因子胁迫引发的、通过氧化跃变产生的信号分子转导,激发细胞程序性死亡机制而表现出的防卫反应.并论述了使该观点成立的依据和进一步研究的策略.该观点的提出,使关于死皮病的各种假说的合理现点达到统一.并对深入研究死皮病的分子机理具有深远的指导意义.     

Polarization optical analysis of blood cell membranes

The present study deals with investigations of membrane structure using polarization topo-optical reactions. Polarization microscopy is a special field of biological submicroscopic morphology. It represents a powerful tool well able to reveal the features of organization of biological structures, and the regularity of macromolecules building cells and tissues - properties that cannot directly be studied by other approaches to complex biological systems. Only in "pure" systems can X-ray diffraction, or the analysis of circular dichroism and the dispersion of optical rotability provide data equivalent to those obtained by polarization microscopy in complex systems. One of the main drawbacks of molecular biology is that most information is relevant to isolated, purified particles or macromolecules. Thus, no conclusions can be drawn concerning the original arrangement of molecules. The gap between biochemical-biophysical and morphological approaches to molecular arrangement in complex structures is bridged by the polarization optical technique. As was pointed out in the introduction, polarization microscopy became a routine biological research method following the pioneering work of Romhányi. His enlightening topo-optical reactions (Romhányi 1960, 1963, 1966) were based on the oriented dye binding of the original charge carriers of regularly arranged tissue constituents. The second group of Romhányi's topo-optical reactions comprised procedures such as sulfation (Romhányi et al. 1973, 1974), the aldehyde-bisulfite-toluidine blue (ABT) reaction (Romhányi et al. 1974, 1975), the permanganate-bisulfite-toluidine blue (PBT) reaction (Fischer 1979, 1979a), and the sialic acid-specific reaction (Makovitzky 1980) all of which operate with induced dye-binding groups; i.e. dye-binding moieties on biological macromolecules are produced by specific chemical reactions.     

NMR spectroscopy reveals unexpected structural variation at the protein–protein interface in MHC class I molecules

It is well established that non-uniform sampling (NUS) allows acquisition of multi-dimensional NMR spectra at a resolution that cannot be obtained with traditional uniform acquisition through the indirect dimensions. However, the impact of NUS on the signal-to-noise ratio (SNR) and sensitivity are less well documented. SNR and sensitivity are essential aspects of NMR experiments as they define the quality and extent of data that can be obtained. This is particularly important for spectroscopy with low concentration samples of biological macromolecules. There are different ways of defining the SNR depending on how to measure the noise, and the distinction between SNR and sensitivity is often not clear. While there are defined procedures for measuring sensitivity with high concentration NMR standards, such as sucrose, there is no clear or generally accepted definition of sensitivity when comparing different acquisition and processing methods for spectra of biological macromolecules with many weak signals close to the level of noise. Here we propose tools for estimating the SNR and sensitivity of NUS spectra with respect to sampling schedule and reconstruction method. We compare uniformly acquired spectra with NUS spectra obtained in the same total measuring time. The time saving obtained when only 1/k of the Nyquist grid points are sampled is used to measure k-fold more scans per increment. We show that judiciously chosen NUS schedules together with suitable reconstruction methods can yield a significant increase of the SNR within the same total measurement time. Furthermore, we propose to define the sensitivity as the probability to detect weak peaks and show that time-equivalent NUS can considerably increase this detection sensitivity. The sensitivity gain increases with the number of NUS indirect dimensions. Thus, well-chosen NUS schedules and reconstruction methods can significantly increase the information content of multidimensional NMR spectra of challenging biological macromolecules.     

Statistical Model for Locating Fragile Points on Nucleic-ACID Molecules

Breakage is an important behavioral characteristic of nucleic-acid molecules. This paper concerns the problem of estimating the location of fragile (F) points on biological macromolecules such as DNA or RNA. We assume that the original length of the macromolecules is a random variable, breakage occurs more likely at some points of the macromolecules than others, and the number of breakages, except for those occurring at such points, is according to a Poisson process. Under these assumptions, the density function of the length of a randomly chosen segment has been derived. The Maximum Likelihood Estimation (MLE) is suggested for estimating the parameters. The model is applied to the data available on RNA of Avian Myeloblostosis Virus (AMV), and three possible F-points have been located. The robustness of the model is discussed.     

A new approach to immunoFET operation

A new method is presented for the detection of an immunological reaction taking place in a membrane, which covers the gate area of an ISFET. By stepwise changing the electrolyte concentration of the sample solution, a transient diffusion of ions through the membrane-protein layer occurs, resulting in a transient membrane potential, which is measured by the ISFET. The diffusion rate is determined by the immobile charge density in the amphoteric protein layer, which changes upon formation of antibody-antigen complexes. No membrane potential is induced at zero fixed charge density as occurs at a protein characteristic pH. Isoelectric points of embedded proteins can be determined by detecting the zero potential response. Up to now, the authors have studied the membrane adsorption of lysozyme, human serum albumin (HSA) and the immune reaction of HSA with the antibody anti-human serum albumin (alpha HSA). The influence of protein parameters on the amplitude of the transient can be described with an empirical equation. Assuming Langmuir behaviour, the protein concentration in the solution can well be correlated with the concentration in the membrane. This new detection method is unique concerning direct measurements of charge densities and isoelectric points of amphoteric macromolecules adsorbed in the membrane. The simple procedure of one incubation stage followed by one detection stage, without separate washing and labelling techniques, gives direct information about specific charge properties of the macromolecules to be studied.     

DNA水凝胶应用于环境样品快速检测的研究进展

DNA作为生物大分子既可以引导生物发育和生命机能活动,也可以被用作构筑纳米生物材料.DNA水凝胶可以制备成兼具DNA生物功能和水凝胶特质,应用于环境样品的分析检测.依据制备DNA水凝胶长链的方法,对比分析了聚合酶链反应、杂交链式反应、滚环扩增技术的制备,物理水凝胶和化学水凝胶的合成过程和改性方法技术特点;并结合环境样品...     

Macromolecular crowding in biological systems: hydrodynamics and NMR methods

Most biologically relevant environments involve highly concentrated macromolecular solutions and most biological processes involve macromolecules that diffuse and interact with other macromolecules. Macromolecular crowding is a general phenomenon that strongly affects the transport properties of macromolecules (rotational and translational diffusion) as well as the position of their equilibria. NMR methods can provide information on molecular interactions, as well as on translational and rotational diffusion. In fact, rotational diffusion, through its determinant role in NMR relaxation, places a practical limit on the systems that can be studied by NMR. While in dilute solutions of non-aggregating macromolecules this limit is set by macromolecular size, in crowded solutions excluded volume effects can have a strong effect on the observed diffusion rates. Hydrodynamic theory offers some insight into the magnitude of crowding effects on NMR observable parameters.     

The temperature jump relaxation technique.

The temperature jump relaxation technique is a convenient and general means of studying rapid reversible reactions of biological macromolecules. Recent advances in automatic data acquisition and the introduction of different optical detection systems will soon allow us to exploit the full potential of kinetic measurements near equilibrium. On the other hand, the kinetic approach can be quite limited if not accompanied by detailed structural and thermodynamic studies. Finally, we must accept the fact that one can rarely demonstrate a reaction mechanism to the exclusion of all plausible alternative models.     

Virus-induced fusion of human erythrocyte ghosts I. Effects of macromolecules on the final stages of the fusion reaction

Human erythrocyte ghosts prepared by hypotonic hemolysis can be fused by Sendai virus, provided that certain macromolecules (bovine serum albumin, dextran and others) are sequestered in the ghosts. Since fusion of the ghosts is dependent on intactness of the F(fusion)-glycoprotein of the virion, and since the other requirements for this reaction are also similar to those for the Sendai virus-induced fusion of intact erythrocytes, this system can be used as a model for the Sendai virus-induced cell fusion reaction. Sequestered macromolecules seem to be required for rounding of locally fused ghosts. Under low osmotic swelling conditions, such as use of ghosts sealed without macromolecules or using bovine serum albumin-loaded ghosts sealed in the presence of external macromolecules, no apparently complete cell fusion (large spherical polyghost formation) could be observed. Even under these conditions, however, occurence of local cell fusion could be demonstrated either by transfer of fluorescent-labeled albumin from one ghost to an other, or by observation of polyghost formation after osmotic swelling in the cold. Thus, final stages of the fusion reaction can be divided into local cell-cell fusion which could not be observed by phase-contrast microscopy, and rounding (i.e. formation of spherical polyghost). For the observation of fusion of ghosts, the last step seems to be important.     

Phase diagrams: a graphical representation of linkage relations

It is shown here that phase diagrams of ligand-binding biological macromolecules provide a powerful tool for the analysis of reaction mechanisms. The present study provides simple rules for the construction and interpretation of such phase diagrams. We give examples for the derivation of reaction schemes for macromolecules that can bind two different kinds of ligands. By sampling one dimension of a phase diagram it is possible to reconstruct the second dimension, including the correct stoichiometry, positive and negative linkage between the ligands and equilibrium binding constants for the complete series of reactions. The discussion is generalised to temperature and pressure-dependent phase diagrams. To exemplify the new diagram method we analyse the pH-dependent binding of trans-beta-indole acrylic acid to apo-Trp repressor, the pH-dependent thermal denaturation of alpha-chymotrypsinogen A, calcium binding and denaturation of annexin I, high affinity zinc binding to a metallo-beta-lactamase and high-pressure and temperature denaturation of RNase A and staphylococcal nuclease.     

Radiation damping compensation of selective pulses in water–protein exchange spectroscopy

The observation of nuclear Overhauser effects (NOEs) between bound water and biological macromolecules such as proteins and nucleic acids can be improved by inverting the water resonance selectively while compensating for radiation damping effects. The efficiency of inversion, the offset profiles, and the appearance of 2D NOE-NOESY spectra can be improved in comparison with earlier methods.