蛋白质复合体及蛋白质相互作用研究新策略—化学交联结合质谱分析法

近年来化学交联法结合质谱分析法被广泛用于蛋白质复合体结构及蛋白质相互作用的研究。研究表明这两种方法的有机结合为研究蛋白质复合体结构及蛋白质相互作用提供了一条新的途径。文章对不同类型的化学交联剂、质谱分析中的Bottom-up 与Top-down 两种研究策略,以及化学交联法结合质谱分析法在蛋白质复合体结构、蛋白质相互作用研究中的应用进行综述。这两种方法的不断发展与完善,将会极大促进生物大分子复合体结构及蛋白质相互作用的研究。     

真核生物转录因子对DNA序列的识别

真核生物转录因子对ＤＮＡ序列的识别杨岐生（浙江大学生物科学与技术系,杭州３１００２７）关键词真核生物转录因子,蛋白质－ＤＮＡ识别研究蛋白质和ＤＮＡ两类生物大分子的相互作用,以阐明基因表达、调控及信息传递的分子机制,是认识生命活动本质的核心问题。本文介...     

蛋白质/核酸相互作用研究方法进展

蛋白质和核酸是构成生命体最为重要的两类生物大分子,蛋白质与核酸的相互作用是分子生物学研究的中心问题之一,它是许多生命活动的重要组成部分。研究蛋白质/核酸相互作用近期采用的新技术有:核酸适体技术、生物信息学方法、蛋白质芯片技术以及纳米技术等。本文就这些新的研究方法进行综述。     

蛋白质相互作用数据库北大核心CSCD

随着"蛋白质组学"的蓬勃发展和人类对生物大分子功能机制的知识积累,涌现出海量的蛋白质相互作用数据。随之,研究者开发了300多个蛋白质相互作用数据库,用于存储、展示和数据的重利用。蛋白质相互作用数据库是系统生物学、分子生物学和临床药物研究的宝贵资源。本文将数据库分为3类:(1)综合蛋白质相互作用数据库;(2)特定物种的蛋白质相互作用数据库;(3)生物学通路数据库。重点介绍常用的蛋白质相互作用数据库,包括BioGRID、STRING、IntAct、MINT、DIP、IMEx、HPRD、Reactome和KEGG等。     

蛋白质相互作用数据库

随着“蛋白质组学”的蓬勃发展和人类对生物大分子功能机制的知识积累,涌现出海量的蛋白质相互作用数据。随之,研究者开发了300多个蛋白质相互作用数据库,用于存储、展示和数据的重利用。蛋白质相互作用数据库是系统生物学、分子生物学和临床药物研究的宝贵资源。本文将数据库分为3类：（1）综合蛋白质相互作用数据库;（2）特定物种的蛋白质相互作用数据库;（3）生物学通路数据库。重点介绍常用的蛋白质相互作用数据库,包括BioGRID、STRING、IntAct、MINT、DIP、IMEx、HPRD、Reactome和KEGG等。     

激光拉曼光谱在蛋白质构象研究中的应用和进展

激光拉曼光谱法被公认为是研究生物大分子的结构、动力学和功能的有效方法。近年来拉曼光谱在蛋白质构象研究中的最新进展,涉及到拉曼光谱在非折叠蛋白质、蛋白质装配的特征描述,拉曼晶体学在实时监控蛋白质单晶中化学变化等方面的应用。另外,介绍了蛋白质拉曼光谱分析在生物技术中的应用现状。并对拉曼光谱技术在蛋白质等生物大分子领域中的研究前景做了进一步的展望。     

蛋白质与DNA相互作用的结构研究

蛋白质与核酸相互作用是生命体内两类最重要的生物大分子,它们之间的相互作用是行使细胞功能的关键,例如:基因复制、转录以及蛋白质的表达翻译等。目前有很多基于蛋白质与DNA复合物的结构研究,这些研究表明蛋白质-DNA的相互作用有很多种方式,所以并不能采用某一种规则或者算法来预测蛋白质与DNA的相互作用,本文主要综述了蛋白质与DNA的相互作用的常用数据库以及结构的特点。     

长链非编码RNA在生物体中的调控作用

长链非编码RNA(Long non-coding RNA,lncRNA)的发现是基因组学和分子生物学研究领域的重要进展。lncRNA在生命活动中具有重要的调节功能,其表达紊乱与多种人类疾病的发生发展密切相关。研究表明,几乎所有的调控性lncRNA通过与不同种类的生物大分子,如DNA、RNA和蛋白质发生相互作用而行使其功能。文章概述了lncRNA在表观遗传学水平、转录水平及转录后水平调控基因表达的效应机制,并探讨了lncRNA如何在肿瘤发生和宿主防御过程中行使功能。不同于小分子ncRNA通过碱基互补配对调控靶基因的表达,大多数已鉴定的lncRNA通过调节蛋白质活性或维持蛋白质复合物的完整性发挥其生物学功能。因此,鉴定lncRNA-蛋白质相互作用可能是理解lncRNA功能的首要任务。     

蛋白质—核酸在基因转录过程中的相互作用

基因表达调腔是当今分子生物学研究的中心内容之一。在整个基因表达调控过程中,充满着蛋白质-核酸及蛋白质-蛋白质相互作用的问题,涉及生命现象的最本质内容。因此,生物大分子的研究,特别是蛋白质-核酸相互作用体系的研究,正从根本上全面地推动着生命科学的发展。就连极具应用前景的生物工程研究,其核心和本质问题也是生物大分子的研究。     

生物科学中一个崭露头角的领域：高静压力研究

通过介绍高静压力作用于生物大分子的机制,提出一些有关高压力生物学的基本概念。此外,还介绍了高静压力在研究蛋白质构象,蛋白质－蛋白质、蛋白质－核酸、蛋白质－配基等相互作用,酶活性的调制以及在生物技术中的应用,表明高压力是一很好的研究手段。     

Signal transduction pathways linking polyamines to apoptosis

Summary. Polyamines are important multifunctional cellular components and are classically considered as mediators of cell growth and division. Recently polyamines have been also implicated in cell death. Now it appears that polyamines are bivalent regulators of cellular functions, promoting proliferation or cell death depending on the cell type and on environmental signals. This review draws a picture about the role of polyamines in signalling pathways related to apoptotic cell death and the proposed molecular targets of these polycations at the level of the apoptotic cascade. Solid evidence indicates that polyamines may affect the mitochondrial and postmitochondrial phases of apoptosis, by modulating cytochrome c release from mitochondria and activation of caspases. Recently, polyamines have been also implicated in the regulation of the premitochondrial phase of apoptosis, during which upstream apoptotic signal transduction pathways are activated. The studies reviewed here suggest that polyamines may participate in loops involving interaction with signal transduction pathways and activation/expression of proteins that may control cell death or cell growth.     

Polyamines: essential factors for growth and survival

Polyamines are low molecular weight, aliphatic polycations found in the cells of all living organisms. Due to their positive charges, polyamines bind to macromolecules such as DNA, RNA, and proteins. They are involved in diverse processes, including regulation of gene expression, translation, cell proliferation, modulation of cell signalling, and membrane stabilization. They also modulate the activities of certain sets of ion channels. Because of these multifaceted functions, the homeostasis of polyamines is crucial and is ensured through regulation of biosynthesis, catabolism, and transport. Through isolation of the genes involved in plant polyamine biosynthesis and loss-of-function experiments on the corresponding genes, their essentiality for growth is reconfirmed. Polyamines are also involved in stress responses and diseases in plants, indicating their importance for plant survival. This review summarizes the recent advances in polyamine research in the field of plant science compared with the knowledge obtained in microorganisms and animal systems.     

Elevated levels of polyamines alter chromatin in murine skin and tumors without global changes in nucleosome acetylation

Polyamines affect nucleosome oligomerization and DNA conformation in vitro, yet little information exists regarding the influence of naturally synthesized polyamines on mammalian chromatin. Capitalizing on the relative inefficiency of a moderate ionic strength extraction buffer to dissociate histones, we obtained evidence of altered chromatin in transgenic mice that overexpress ornithine decarboxylase (ODC), which catalyzes polyamine synthesis. Dissociation of histones from chromatin in ODC transgenic mouse skin, as well as in tumors that develop spontaneously in ODC/Ras bigenic mice, is dramatically reduced relative to normal littermate skin. This could reflect tighter tethering of nucleosomes to DNA or a more compacted chromatin structure due to elevated intracellular concentrations of polyamines since this effect is reversible upon treatment with alpha-difluoromethylornithine (DFMO), a specific inhibitor of ODC enzymatic activity. Impeded release of nonhistone chromatin proteins HP-1beta and nucleophosmin, but not Lamin B, HDAC-1, HMGB, HMGN2, or HMGA1, suggests that polyamines exert selective effects on specific chromatin protein complexes. Moreover, overall acetylation, as well as specific methylation, of nucleosomes in ODC mice is unaffected, implying that access by histone modifying enzymes is not generally restricted. The abnormal chromatin environment fostered by elevated levels of polyamines may be a necessary prerequisite for epithelial tumor growth and maintenance.     

Polyamines: mysterious modulators of cellular functions

In recent years the functions of polyamines (putrescine, spermidine, and spermine) have been studied at the molecular level. Polyamines can modulate the functions of RNA, DNA, nucleotide triphosphates, proteins, and other acidic substances. A major part of the cellular functions of polyamines can be explained through a structural change of RNA which occurs at physiological concentrations of Mg(2+) and K(+) because most polyamines exist in a polyamine-RNA complex within cells. Polyamines were found to modulate protein synthesis at several different levels including stimulation of special kinds of protein synthesis, stimulation of the assembly of 30 S ribosomal subunits and stimulation of Ile-tRNA formation. Effects of polyamines on ion channels have also been reported and are gradually being clarified at the molecular level.     

Role of polyamines in the ontogeny of plants and their biotechnological applications

Recent developments in the metabolism and function of polyamines in plants is presented. Polyamines appear to be involved in a wide range of plant processes, however their exact role is not completely understood. In this review, the metabolic pathways involved in polyamine biosynthesis and degradation are explained, along with the transport and conjugation of these compounds. The studies involved in the understanding of function(s) of polyamines using metabolic inhibitors, as well as genetic and molecular approaches are described. Polyamine metabolism and profound changes in polyamine titres in response to infection by pathogens has been presented. Its role in adaptation of plants to stress is also presented. Molecular understanding of polyamines and their modulation in transgenics is also discussed. Further line of work in the understanding of the role of polyamines has also been focussed.     

Polyamines alter sequence-specific DNA-protein interactions.

The polyamines are abundant biogenic cations implicated in many biological processes. Despite a plethora of evidence on polyamine-induced DNA conformational changes, no thorough study of their effects on the activities of sequence-specific DNA binding proteins has been performed. We describe the in vitro effects of polyamines on the activities of purified, representative DNA-binding proteins, and on complex protein mixtures. Polyamines at physiological concentrations enhance the binding of several proteins to DNA (e.g. USF, TFE3, Ig/EBP, NF-IL6, YY1 and ICP-4, a herpes simplex virus gene regulator), but inhibit others (e.g. Oct-1). The degree of enhancement correlates with cationic charge; divalent putrescine is ineffective whereas tetravalent spermine is more potent than trivalent spermidine. Polyamine effects on USF and ICP-4 result from increased rate of complex formation rather than a decreased rate of dissociation. DNAse I footprint analysis indicated that polyamines do not alter DNA-protein contacts. Polyamines also facilitate formation of complexes involving binding of more than one protein on a DNA fragment.     

Polyamines alter the phosphorylation pattern of chromatin proteins by endogenous protein kinases

The effect of polyamines on the chromatin phosphorylation by endogenous protein kinases was investigated. Polyamines not only selectively stimulated the phosphorylation of chromatin proteins but also concurrently inhibited the phosphorylation of a number of polypeptides. In particular, a 11,000-dalton polypeptide with pI 4.5–5.0 was highly phosphorylated in the absence of polyamines, despite being a minor component whereas the phosphorylation was strongly inhibited in the presence of polyamines.     

Polyamines and mRNA stability in regulation of intestinal mucosal growth

Summary. The mammalian intestinal epithelium is a rapidly self-renewing tissue in the body, and its homeostasis is preserved through strict regulation of epithelial cell proliferation, growth arrest, and apoptosis. Polyamines are necessary for normal intestinal mucosal growth and decreasing cellular polyamines inhibits cell proliferation and disrupts epithelial integrity. An increasing body of evidence indicates that polyamines regulate intestinal epithelial cell renewal by virtue of their ability to modulate expression of various genes and that growth inhibition following polyamine depletion results primarily from the activation of growth-inhibiting genes rather than a simple decrease in expression of growth-promoting genes. In this review article, we will focus on changes in expression of growth-inhibiting genes following polyamine depletion and further analyze in some detail the mechanisms through which mRNA stability is regulated by RNA-binding proteins.