生物质谱技术与蛋白质组学

生物质谱技术是蛋白质组学的支撑技术.详细论述了质谱技术的分类与基本分析原理,重点论述了质谱技术的发展变化,包括基质辅助激光解吸飞行时间质谱技术,电喷雾质谱技术,MALDI-Q-TOF和MAL-DI-TOF-TOF等质谱技术,以及质谱技术在蛋白质组学研究中的应用与未来的发展和挑战.     

生物质谱技术在蛋白质组学研究中的应用

随着技术的进步,蛋白质组学的研究重心由最初旨在鉴定细胞或组织内基因组所表达的全部蛋白质转移到从整个蛋白质组水平上阐述包括蛋白翻译后修饰、生物大分子相互作用等反映蛋白质功能的层次。多种质谱离子化技术的突破使质谱技术成为蛋白质组学研究必不可少的手段。质谱技术联合蛋白质组学多角度、深层次探索生命系统分子本质成为现阶段生命科学研究领域的主旋律之一。本文简要综述了肽和蛋白质等生物大分子质谱分析的原理、方式和应用,并对其发展前景做出展望。     

Solid-phase Chemistry: A Useful Tool to Discover Modulators of Protein Interactions

The Solid phase synthesis (SPS) concept, first developed for biopolymers, has spread in every field where organic synthesis is involved. While the potential of the solid-phase method was obvious in 1959 to its discoverer, Prof. R. B. Merrifield, it was unpredictable its dominance in peptide synthesis and especially in combinatorial chemistry, an area not yet conceived. SPS paved the way for solid-phase combinatorial approaches (extensively reviewed in (Choong, I. C. and Ellman, J. A.: 1996, Annu. Rep. Med. Chem. 31, 309–318; Obrecht, D. and Villalgordo, J. M.: 1998, Solid-supported Combinatorial and Parallel Synthesis of Small-Molecular-Weight Compound Libraries. Pergamon Press Ltd., Oxford, UK; Chabala, J. C.: 1995, Curr. Opin. Biotechnol. 6, 632–639; Kamal, A., Reddy, K. L., Devaiah, V., Shankaraiah, N., Reddy, D. R.: 2006, Mini Rev. Med. Chem. 6, 53–69; Whitehead, D. M., McKeown, S. C., Routledge, A.: 2005, Comb. Chem. HTS 8, 361–371; Nefzi, A., Ostresh, J. M., Houghten, R. A.: 1997, Chem. Rev. 97, 449–472; Gordon, E. M., Gallop, M. A., Patel, D. V.: 1996, Acc. Chem. Res. 29, 144–154)) as many laboratories and companies focused on the development of technologies and chemistry suitable to this new methodology. This resulted in the spectacular outburst of combinatorial chemistry, which profoundly changed the approach for new drug discovery. Combinatorial chemistry is currently considered a valid approach for a wide range of biomedical applications, such as, target validation and drug discovery.     

Proteomics approaches to biomarker detection.

The development of mass spectrometry (MS) technologies has brought the ability to gather massive amounts of data characterising the proteomes of complex mixtures. A major focus in proteomics is to leverage this data-gathering capability to conduct comparative analyses of biofluids from healthy and disease-affected patients for the identification of highly specific biomarkers and/or the development of MS-based diagnostic platforms. Much effort has gone into optimising the biofluid proteome coverage that can be obtained using these technologies, leaving proteomics poised to make an important impact in disease diagnostics in the future.     

细胞质膜蛋白质组学研究技术进展

质膜蛋白在细胞中执行着非常重要的功能。随着蛋白质组学的发展,细胞质膜蛋白质组学成为蛋白质组学研究的重要组成部分,它为质膜蛋白的生物功能研究及药物靶标的发现提供了新的途径。然而,质膜蛋白丰度低、疏水性强,对现有蛋白质组学研究技术提出了挑战。简要综述了近年来质膜蛋白质组研究的相关技术进展,包括富集、提取分离鉴定方法及定量和生物信息学研究方法等。     

高通量植物蛋白质组学研究方法

模式植物拟南芥和水稻的基因组测序,使得大规模、高通量的研究方法在基因组和蛋白质组研究中日趋重要。本文综述双向电泳、质谱、蛋白质微阵列、抗体、酵母双杂交系统以及一些新型高通量方法研究进展及其在植物蛋白质组研究中的应用。     

Proteomics and a future generation of plant molecular biologists

Proteomic methods are required for the study of many different aspects of plant function. Important issues in proteomics include the molecular complexity of proteins, given that there are hundreds of thousands of chemically and physically distinct proteins in plants, and the context of protein functions with respect to both genomes and the environment. Available genomic and gene sequences greatly simplify the identification of proteins using improved techniques of mass spectrometry. This improved capability has led to much discussion on proteomes, and some experimentation using proteomic methodologies aimed at modest numbers of proteins. The scale of proteomics is open, for the number of proteins and genes considered at any one time is as dependent on the nature of the scientific question posed as on technical resources and capabilities. We know just enough about plant proteomes to imagine the breathtaking scope of our ignorance. There are tremendous opportunities for new molecular biologists to define the nature of the protein machines that transduce genetic and environmental information, and transform simple energy and matter, to give plants.     

Chemical proteomics and its impact on the drug discovery process

Despite the rapid growth of postgenomic data and fast-paced technology advancement, drug discovery is still a lengthy and difficult process. More effective drug design requires a better understanding of the interaction between drug candidates and their targets/off-targets in various situations. The ability of chemical proteomics to integrate a multiplicity of disciplines enables the direct analysis of protein activities on a proteome-wide scale, which has enormous potential to facilitate drug target elucidation and lead drug verification. Over recent years, chemical proteomics has experienced rapid growth and provided a valuable method for drug target identification and inhibitor discovery. This review introduces basic concepts and technologies of different popular chemical proteomic approaches. It also covers the essential features and recent advances of each approach while underscoring their potentials in drug discovery and development.     

定量蛋白质组同位素标记技术及应用

定量蛋白质组学是对蛋白质组进行精确的定量和鉴定的学科,突破了传统蛋白质组研究集中于对蛋白质的分离和鉴定,着重于定性定量解析细胞蛋白质的动态变化信息,更真实地反映了细胞功能、过程机制等综合信息。以同位素为内标的质谱分析新技术的提出,显示出可同时自动鉴定和精确定量的能力,代表了目前定量蛋白质组研究的主要发展方向。对近年来定量蛋白质组学同位素标记技术和应用研究所取得的重要进展以及最新的发展动态进行了综述。     

Proteomics and metabolomics in cancer drug development

In this review article, the main recent advancements in the field of proteomics and metabolomics and their application in cancer research are described. In the second part of the review the main metabolic alterations observed in cancer cells are thoroughly dissected, especially those involving anabolic pathways and NADPH-generating pathways, which indirectly affect anabolic reactions, other than the maintenance of the redox poise. Alterations to mitochondrial pathways and thereby deriving oncometabolites are also detailed. The third section of the review is a discussion of how and to what extent (mutations to) tumor suppressors and oncogenes end up influencing cancer cell metabolism and cell fate, either promoting survival and proliferation or autophagy and apoptosis. In the last section of the review, an overview is provided of therapeutic strategies that make use of metabolic reprogramming approaches.     

蛋白质组学研究中的一种新方法——配体垂钓

基于表面等离子共振技术的配体垂钓技术能在蛋白质组水平上研究蛋白质的相互作用与功能,提供控制细胞功能的新靶标．其通过将受体固定在芯片表面,当被检测样品流过芯片表面时,配体与受体相结合, 实现俘获未知的相互作用的伙伴蛋白或复合体,并结合质谱技术鉴定出未知蛋白及其序列．     

Characterization of Protomer Interfaces in HslV Protease; the Bacterial Homologue of 20S Proteasome

HslVU, a two-component proteasome-related prokaryotic system is composed of HslV protease and HslU ATPase. HslV protomers assemble in a dodecamer of two-stacked hexameric rings that form a complex with HslU hexamers. The intra- and inter-ring protomer interfaces in the HslV dodecamer underpin the integrity and functionality of HslVU. Structural characterization of HslV from different bacteria illustrated considerable differences in interacting residues, accessible surface and gap volumes at the intra-ring interface that is primarily stabilized by polar interactions. Amino acid residues Lys28, Arg83 and Asp111 have envisaged as hot spots at this HslU-interacting interface. The inter-ring interfaces that are made up of side chain packing of hydrophobic residues are structurally conserved. Hyperthermostable bacterium T. maritima HslV has extensively networked polar/nonpolar interactions and highly packed environment at all interfaces. Present data demonstrates that HslV protomer interfaces perform distinct functions; whereas intra-ring interface participates in HslV:HslU interaction resulting in allosteric activation of HslV protease by HslU, the inter-ring interfaces uphold the oligomeric form of HslV.     

Proteomics in atherothrombosis: a future perspective

Atherothrombosis is the primary cause of death in Western countries. The cellular and molecular mechanisms underlying atherosclerosis remain widely unknown. The complex nature of atherosclerotic cardiovascular diseases demands the development of novel technologies that enable discovery of new biomarkers for early disease detection and risk stratification, which may predict clinical outcome. In this review, we outline potential sources and recent proteomic approaches that could be applied in the search of novel biomarkers of cardiovascular risk. In addition, we describe some issues raised in relation to the application of proteomics to blood samples, as well as two novel emerging concepts, such as peptidomics and population proteomics. In the future, the use of high-throughput techniques (proteomic, genomics and metabolomics) will potentially identify novel patterns of biomarkers, which, along with traditional risk factors and imaging techniques, could help to target vulnerable patients and monitor the beneficial effects of pharmacological agents.     

An atlas of protein-protein interactions across mouse tissues

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Activity-based proteomics: enzymatic activity profiling in complex proteomes

Summary. In the postgenomic era new technologies are emerging for global analysis of protein function. The introduction of active site-directed chemical probes for enzymatic activity profiling in complex mixtures, known as activity-based proteomics has greatly accelerated functional annotation of proteins. Here we review probe design for different enzyme classes including serine hydrolases, cysteine proteases, tyrosine phosphatases, glycosidases, and others. These probes are usually detected by their fluorescent, radioactive or affinity tags and their protein targets are analyzed using established proteomics techniques. Recent developments, such as the design of probes for in vivo analysis of proteomes, as well as microarray technologies for higher throughput screenings of protein specificity and the application of activity-based probes for drug screening are highlighted. We focus on biological applications of activity-based probes for target and inhibitor discovery and discuss challenges for future development of this field.