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1.
Peptide aptamers are peptides constrained and presented by a scaffold protein that are used to study protein function in cells. They are able to disrupt protein-protein interactions and to constitute recognition modules that allow the creation of a molecular toolkit for the intracellular analysis of protein function. The success of peptide aptamer technology is critically dependent on the performance of the scaffold. Here, we describe a rational approach to the design of a new peptide aptamer scaffold. We outline the qualities that an ideal scaffold would need to possess to be broadly useful for in vitro and in vivo studies and apply these criteria to the design of a new scaffold, called STM. Starting from the small, stable intracellular protease inhibitor stefin A, we have engineered a biologically neutral scaffold that retains the stable conformation of the parent protein. We show that STM is able to present peptides that bind to targets of interest, both in the context of known interactors and in library screens. Molecular tools based on our scaffold are likely to be used in a wide range of studies of biological pathways, and in the validation of drug targets. 相似文献
2.
Cancer is a highly complex and heterogeneous disease involving a succession of genetic changes (frequently caused or accompanied by exogenous trauma), and resulting in a molecular phenotype that in turn results in a malignant specification. The development of malignancy has been described as a multistep process involving self-sufficiency in growth signals, insensitivity to antigrowth signals, evasion of apoptosis, limitless replicative potential, sustained angiogenesis, and finally tissue invasion and metastasis. The quantitative analysis of networking molecules within the cells might be applied to understand native-state tissue signalling biology, complex drug actions and dysfunctional signalling in transformed cells, that is, in cancer cells. High-content and high-throughput single-cell analysis can lead to systems biology and cytomics. The application of cytomics in cancer research and diagnostics is very broad, ranging from the better understanding of the tumour cell biology to the identification of residual tumour cells after treatment, to drug discovery. The ultimate goal is to pinpoint in detail these processes on the molecular, cellular and tissue level. A comprehensive knowledge of these will require tissue analysis, which is multiplex and functional; thus, vast amounts of data are being collected from current genomic and proteomic platforms for integration and interpretation as well as for new varieties of updated cytomics technology. This overview will briefly highlight the most important aspects of this continuously developing field. 相似文献
3.
合成生物学是一个基于生物学和工程学原理的科学领域,其目的是重新设计和重组微生物,以优化或创建具有增强功能的新生物系统。该领域利用分子工具、系统生物学和遗传框架的重编程,从而构建合成途径以获得具有替代功能的微生物。传统上,合成生物学方法通常旨在开发具有成本效益的微生物细胞工厂进而从可再生资源中生产化学物质。然而,近年来合成生物学技术开始在环境保护中发挥着更直接的作用。本综述介绍了基因工程中的合成生物学工具,讨论了基于基因工程的微生物修复策略,强调了合成生物学技术可以通过响应特定污染物进行生物修复来保护环境。其中,规律间隔成簇短回文重复序列(Clustered Regularly Interspersed Short Palindromic Repeats, CRISPR)技术在基因工程细菌和古细菌的生物修复中得到了广泛应用,生物修复领域也出现了很多新的先进技术,包括生物膜工程、人工微生物群落的构建、基因驱动、酶和蛋白质工程等。有了这些新的技术和工具,生物修复将成为当今最好和最有效的污染物去除方式之一。 相似文献
4.
Weiqiang Li Jie Qin Xinyu Li Li Zhang Chang Liu Fei Chen Zifei Wang Lirong Zhang Xiuming Zhang Bruce T. Lahn Andy Peng Xiang 《生物学前沿》2010,5(5):455-463
The ability of human embryonic stem cells (hESCs)to undergo indefinite self-renewal in vitro and to produce lineages derived from all three embryonic germ layers both in vitro and in vivo makes such cells extremely valuable in both clinical and research settings.However,the generation of specialized cell lineages from a mixture of differentiated hESCs remains technically difficult.Tissue specific promoter-driven reporter genes are powerful tools for tracking cell types of interest in differentiated cell populations.Here,we describc the construction of modular lentivectors containing different tissue-specific promoters(Tαl of α-tubulin:αP2 of adipocyte Protein 2;and AFP of alpha fetoprotein)driving expression of humanized Renilla green fluorescent protein(hrGFP).To this end,we used MultiSite gateway technology and employed the novel vectors to successfully monitor hESC differentiation.We present a versatile method permitting target cells to bc traced.Our system will facilitate research in developmental biology,transplantation,and in vivo stem cell tracking. 相似文献
5.
The 'right' size in nanobiotechnology 总被引:8,自引:0,他引:8
Whitesides GM 《Nature biotechnology》2003,21(10):1161-1165
The biological and physical sciences share a common interest in small structures (the definition of 'small' depends on the application, but can range from 1 nm to 1 mm). A vigorous trade across the borders of these areas of science is developing around new materials and tools (largely from the physical sciences) and new phenomena (largely from the biological sciences). The physical sciences offer tools for synthesis and fabrication of devices for measuring the characteristics of cells and sub-cellular components, and of materials useful in cell and molecular biology; biology offers a window into the most sophisticated collection of functional nanostructures that exists. 相似文献
6.
Adaptive Profiling (APL) and other biochip companies aim to harness the power of microsystems technology together with advances in chemistry and molecular biology, to become service and technology providers to organizations involved in pharmaceutical research and development. By supplying a unique range of decision-making tools that aid an earlier identification of qualified drug candidates for clinical development, the company should gain a significant share of the 10 billion US dollar biological screening, bioavailability and toxicity assessment market. 相似文献
7.
A low-volume platform for cell-respirometric screening based on quenched-luminescence oxygen sensing
Alderman J Hynes J Floyd SM Krüger J O'Connor R Papkovsky DB 《Biosensors & bioelectronics》2004,19(11):1529-1535
Cell viability assays represent an important technology in modern cell biology, drug discovery and biotechnology, where currently there is a high demand for simple, sensitive and cost-effective screening methods. We have developed a new methodology and associated tools for cell-based screening assays, which are based on the measurement of the rates of oxygen uptake in cells by luminescence quenching. Sealable microchamber devices matching the footprint of a standard 96-well plate were developed and used in conjunction with long-decay phosphorescent oxygen probes. These devices permit cell non-invasive, real-time monitoring of cellular respiration and a rapid, one-step, kinetic assessment of multiple samples for cell viability, drug/effector action. These assays can be carried out on conventional fluorescence plate readers, they are suitable for different types of cells, including adherent and slow-respiring cells, require small sample volumes and cell numbers, and are amenable for high throughput screening. Monitoring of as little as 300 mammalian cells in 3 microl volume has been demonstrated. 相似文献
8.
Baculovirus display strategies: Emerging tools for eukaryotic libraries and gene delivery. 总被引:3,自引:0,他引:3
Christian Oker-Blom Kari J Airenne Reingard Grabherr 《Briefings in Functional Genomics and Prot》2003,2(3):244-253
Recombinant baculoviruses have been extensively used as vectors for abundant expression of a large variety of foreign proteins in insect cell cultures. The appeal of the system lies essentially in easy cloning techniques and virus propagation combined with the eukaryotic post-translational modification machinery of the insect cell. Recently, a novel molecular biology tool was established by the development of baculovirus surface display, using different strategies for presentation of foreign peptides and proteins on the surface of budded virions. This eukaryotic display system enables presentation of large complex proteins on the surface of baculovirus particles and has thereby become a versatile system in molecular biology. Surface display strategies play an important role, as they may be used to enhance the efficiency and specificity of viral binding and entry to mammalian cells. In addition, baculovirus surface display vectors have been engineered to contain mammalian promoter elements designed for gene delivery both in vitro and in vivo. Moreover, baculovirus capsid display has recently been developed; this holds promise for intracellular targeting of the viral capsid and subsequent cytosolic delivery of desired protein moieties. Finally, the viruses can accommodate large insertions of foreign DNA and replicate only in insect cells. Together, these are attributes that are very likely to make them important tools in functional genomics and proteomics. 相似文献
9.
In recent years, peptide aptamers have emerged as novel molecular tools that are useful for both basic and applied aspects of molecular medicine. Due to their ability to specifically bind to and inactivate a given target protein at the intracellular level, they provide a new experimental strategy for functional protein analyses, both in vitro and in vivo. In addition, by using peptide aptamers as "pertubagens", they can be employed for genetic analyses, in order to identify biochemical pathways, and their components, that are associated with the induction of distinct cellular phenotypes. Furthermore, peptide aptamers may be developed into diagnostic tools for the detection of a given target protein or for the generation of high-throughput protein arrays. Finally, the peptide aptamer technology has direct therapeutic implications. Peptide aptamers can be used in order to validate therapeutic targets at the intracellular level. Moreover, the peptide aptamer molecules themselves should possess therapeutic potential, both as lead structures for drug design and as a basis for the development of protein drugs. 相似文献
10.
The study of mechanisms that underlie Parkinson's disease (PD), as well as translational drug development, has been hindered by the lack of appropriate models. Both cell culture systems and animal models have limitations, and to date none faithfully recapitulate all of the clinical and pathological phenotypes of the disease. In this review we examine the various cell culture model systems of PD, with a focus on different stem cell models that can be used for investigating disease mechanisms as well as drug discovery for PD. We conclude with a discussion of recent discoveries in the field of stem cell biology that have led to the ability to reprogram somatic cells to a pluripotent state via the use of a combination of genetic factors; these reprogrammed cells are termed “induced pluripotent stem cells” (iPSCs). This groundbreaking technique allows for the derivation of patient-specific cell lines from individuals with sporadic forms of PD and also those with known disease-causing mutations. Such cell lines have the potential to serve as a human cellular model of neurodegeneration and PD when differentiated into dopaminergic neurons. The hope is that these iPSC-derived dopaminergic neurons can be used to replicate the key molecular aspects of neural degeneration associated with PD. If so, this approach could lead to transformative new tools for the study of disease mechanisms. In addition, such cell lines can be potentially used for high-throughput drug screening. While not the focus of this review, ultimately it is envisioned that techniques for reprogramming of somatic cells may be optimized to a point sufficient to provide potential new avenues for stem cell-based restorative therapies. 相似文献
11.
Ernest Giralt 《Journal of peptide science》2015,21(6):447-453
Peptides are destined to play a major role as therapeutic agents. My laboratory is contributing to speeding up this process. On the one hand, we devote efforts to studying the molecular details and dynamics of the events that occur during molecular recognition at protein surfaces. We succeeded to design and synthesize peptides able to modulate these recognition events either permanently or in response to light. On the other hand, we are discovering and designing peptides able to cross biological barriers. Our aim is to use these peptides as shuttles for targeting therapeutic agents to organs, tissues, or cells, with a special emphasis on drug delivery to the brain. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
12.
The flow of cytometry into systems biology. 总被引:1,自引:0,他引:1
Biomedical research is evolving to address biological systems as molecular pathways integrated into complex networks. Tools for molecular and cell analysis are also evolving to address the new challenges and opportunities of this approach. Flow cytometry is a versatile analytical platform, capable of high speed quantitative measurements of cells and other particles. These capabilities are being exploited and extended in a range of new applications stemming from opportunities presented by the advances of genomics, proteomics and systems biology, which are in turn impacting clinical diagnosis, vaccine development and drug discovery. In this review, we highlight some of these advances and consider the future evolution of flow cytometry technology. 相似文献
13.
多能干细胞(pluripotent stem cell,PSC)是一类具有自我更新能力和多向分化潜能的细胞,具有广泛的临床应用前景.诱导性多功能干细胞(induced pluripotent stem cell,iPS cell)的获得,解决了传统方式中的细胞来源和伦理学等问题,从理论研究和应用上实现了体细胞重编程的重大突破,也为疾病发生机制研究、药物筛选、个性化药物选择、细胞治疗和再生医学等研究创造了难得的机会,从而开启了多能干细胞应用的新纪元.iPS过程中有很多问题尚未得到解决,尤其是诱导重编程的分子机制方面,这也是近年来干细胞领域研究的热点.其中如何实现表观遗传的重编程被认为是亟待解决的核心问题之一.本文结合我们的研究,主要介绍诱导重编程领域表观遗传修饰重塑机制的研究进展,并展望未来研究中大规模信息整合分析的重要性. 相似文献
14.
The scientific techniques used in molecular biological research and drug discovery have changed dramatically over the past 10 years due to the influence of genomics, proteomics and bioinformatics. Furthermore, genomics and functional genomics are now merging into a new scientific approach called chemogenomics. Advancements in the study of molecular cell biology are dependent upon "omics" researchers realizing the importance of and using the experimental tools currently available to cell biologists. For example, novel microscopic techniques utilizing advanced computer imaging allow for the examination of live specimens in a fourth dimension, viz., time. Yet, molecular biologists have not taken full advantage of these and other traditional and novel cell biology techniques for the further advancement of genomic and proteomic-oriented research. The application of traditional and novel cellular biological techniques will enhance the science of genomics. The authors hypothesize that a stronger interdisciplinary approach must be taken between cell biology (and its closely related fields) and genomics, proteomics and bio-chemoinformatics. Since there is a lot of confusion regarding many of the "omics" definitions, this article also clarifies some of the basic terminology used in genomics, and related fields. It also reviews the current status and future potential of chemogenomics and its relationship to cell biology. The authors also discuss and expand upon the differences between chemogenomics and the relatively new term--chemoproteomics. We conclude that the advances in cell biology methods and approaches and their adoption by "omics" researchers will allow scientists to maximize our knowledge about life. 相似文献
15.
16.
Thomas C. G. Bosch Prof. Dr. Dr. h.c. 《当今生物学》2009,39(2):114-122
The basis for Hydra's enormous regeneration capacity and potential immortality is a life cycle in which proliferation occurs mostly asexual by budding. That requires that each polyp contains a large number of cells which continuously undergo self‐renewing mitotic divisions and also have the option to follow differentiation pathways. Now, emerging molecular tools for the first time shed light on the molecular processes controlling these pathways. Studies of stem cells in Hydra, therefore, promise critical insights of general relevance into stem cell biology and the evolutionary origin of these cells. 相似文献
17.
Cell culture methods and models are key investigative tools for cell and molecular biology studies. Fetal bovine serum (FBS) is commonly used as an additive during cell culture since its constituents promote cell survival, proliferation and differentiation. Here we report that commercially available FBS from different major suppliers consistently contain precipitated, calcium oxalate crystals-either in the monohydrate (COM) or dihydrate (COD) form. Mineral structure and phase identification of the crystals were determined by X-ray diffraction, chemical composition by energy-dispersive X-ray microanalysis, and imaging and measurement of crystal growth steps by atomic force microscopy-all identified and confirmed crystallographic parameters for COM and COD. Proteins binding to the crystals were identified by immunoblotting, revealing the presence of osteopontin and fetuin-A (alpha(2)HS-glycoprotein)--known regulators of crystal growth found in serum. Macrophage cell cultures exposed to calcium oxalate crystals showed internalization of the crystals by phagocytosis in a process that induced disruption of cell-cell adhesion, release of reactive oxygen species and membrane damage, events that may be linked to the release of inflammatory cytokines by these cells into the culture media. In conclusion, calcium oxalate crystals found in commercially available FBS are toxic to cells, and their presence may confound results from in vitro studies where, amongst others, phagocytosis, biomineralization, renal cell and molecular biology, and drug and biomaterial testing are being examined. 相似文献
18.
Expanded fluorescent protein techniques employing photo-switchable and
fluorescent timer proteins have become important tools in biological research.
These tools allow researchers to address a major challenge in cell and
developmental biology, namely obtaining kinetic information about the processes
that determine the distribution and abundance of proteins in cells and tissues.
This knowledge is often essential for the comprehensive understanding of a
biological process, and/or required to determine the precise point of
interference following an experimental perturbation. 相似文献
19.
20.
Wallace L. Mc Keehan David Barnes Lola Reid Eric Stanbridge Hiroki Murakami Gordon H. Sato 《In vitro cellular & developmental biology. Plant》1990,26(1):9-23
Summary For the past 60 years, fundamental discoveries in eukaryotic biology using mammalian cell cultures have been significant but
modest relative to the enormous potential. Combined with advances in technologies of cell and molecular biology, mammalian
cell culture technology is becoming a major, if not essential tool, for fundamental discovery in eukaryotic biology. Reconstruction
of the milieu for cells has progressed from simple salt solutions supporting brief survival of tissues outside the body to
synthesis of the complete set of structurally defined nutrients, hormones and elements of the extracellular matrix needed
to reconstruct complex tissues from cells. The isolation of specific cell types in completely defined environments reveals
the true complexity of the mammalian cell and its environment as a dynamic interactive physiological unit. Cell cultures provide
the tool for detection and dissection of the mechanism of action of cellular regulators and the genes that determine individual
aspects of cell behavior. The technology underpins advances in virology, somatic cell genetics, endocrinology, carcinogenesis,
toxicology, pharmacology, hematopoiesis and immunology, and is becoming a major tool in develomental biology, complex tissue
physiology and production of unique mammalian cell-derived biologicals in industry.
This article is the first of a series of invited reviews aimed at identifying fundamental contributions and current challenges
associated with research activities in subdiscriplines of cell and developmental biology in vitro. This treatise is dedicated
to Dr. Brian Kimes, Program Director at the National Cancer Institute, whose vision, encouragement and support have contributed
significantly to modern developments in mammalian cell culture. 相似文献