Abstract：
:Use of fluorescence in biology and biochemistry for imaging and characterizing equilibrium and dynamic processes is growing exponentially. Much progress has been made in the last few years on the microscopic understanding of the underlying principles of what controls the wavelength and quenching of fluorescence in biopolymers, both of which are central to the utility of fluorescent probes. This chapter is concerned with the quantitative microscopic understanding and prediction of the fluorescence wavelength and/or intensity of a fluorescent probe molecule attached to a biopolymer as revealed by hybrid quantum and classical mechanical computation procedures. The aim is not only to provide a recipe, but also even more importantly, to communicate the qualitative basic concepts of interpretation of fluorescence. These are surprisingly simple, although not broadly appreciated at this time. In addition, an effort has been made to show how these techniques have led to an emerging understanding of the relation between time-dependent wavelengths shifts due to solvent relaxation and population decay of conformational sub-ensembles.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
Callis PRdoi
10.1016/B978-0-12-381270-4.00001-9subject
Has Abstractpub_date
2011-01-01 00:00:00pages
1-38eissn
0076-6879issn
1557-7988pii
B978-0-12-381270-4.00001-9journal_volume
487pub_type
杂志文章abstract：:One of the greatest advances in molecular genetics has been the application of selectable transposons in molecular biology. After 30 years of use in microbial genetics studies, transposons remain indispensable tools for the generation of null alleles tagged with selectable markers, genetic mapping, manipulation of chr...
journal_title：Methods in enzymology
pub_type： 杂志文章,评审
doi：10.1016/S0076-6879(06)21002-4
更新日期：2007-01-01 00:00:00
abstract：:Hybrid proteins or chimeras are generated by recombination of protein fragments. In the course of evolution, this mechanism has led to major diversification of protein folds and their functionalities. Similarly, protein engineers have taken advantage of this attractive strategy to build new proteins. Methods that use ...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-394292-0.00018-7
更新日期：2013-01-01 00:00:00
abstract：:The nematode Caenorhabditis elegans has served as a fruitful setting for cell death research for over three decades. A conserved pathway of four genes, egl-1/BH3-only, ced-9/Bcl-2, ced-4/Apaf-1, and ced-3/caspase, coordinates most developmental cell deaths in C. elegans. However, other cell death forms, programmed and...
journal_title：Methods in enzymology
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doi：10.1016/B978-0-12-801430-1.00007-X
更新日期：2014-01-01 00:00:00
abstract：:PqqE is the first enzyme in the biosynthetic pathway of the redox cofactor pyrroloquinoline quinone (PQQ), catalyzing the formation of a carbon-carbon bond in the precursor peptide PqqA. PqqE is a radical S-adenosyl-l-methionine (SAM) (RS) enzyme, a family of enzymes that use the reductive cleavage of a [4Fe-4S] clust...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2018.04.002
更新日期：2018-01-01 00:00:00
abstract：:Determination of the three-dimensional structures of the globins led to the problem of determining how the ligands bound to the heme. In many of these structures there was no clear path from the solvent to the ligand-binding site. Even in those structures that appeared to have one or more tunnels from the exterior to ...
journal_title：Methods in enzymology
pub_type： 杂志文章,评审
doi：10.1016/S0076-6879(07)37021-3
更新日期：2008-01-01 00:00:00
abstract：:A fundamental concept of phosphorylation-mediated signaling is the precise switching between discrete functional conformations. According to the traditional view, phosphorylation induces a new, active conformation. In this chapter, a series of NMR experiments performed on a response regulator are described that challe...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(07)23006-X
更新日期：2007-01-01 00:00:00
abstract：:The characterization of microbial adhesins is greatly facilitated by the use of synthetic peptides. Synthetic peptides can be used to identify specific antigenic epitopes, to delineate receptor-binding domain of adhesins, and to facilitate the characterization of the adhesin, and they allow for a direct examination of...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/s0076-6879(95)53013-4
更新日期：1995-01-01 00:00:00
abstract：:Multiprotein machines drive virtually all primary cellular processes. Modular multidomain proteins are widely distributed within these dynamic complexes because they provide the flexibility needed to remodel structure as well as rapidly assemble and disassemble components of the machinery. Understanding the functional...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2017.03.017
更新日期：2017-01-01 00:00:00
abstract：:Liposomes show great promise as intravenous drug delivery vehicles, but it is often difficult to combine stability in the circulation with rapid, targeted release at the site of interest. Targeting to specific tissues requires developing highly specific ligands with strong affinities to receptors overexpressed on dise...
journal_title：Methods in enzymology
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doi：10.1016/S0076-6879(09)64014-3
更新日期：2009-01-01 00:00:00
abstract：:Two-component signal transduction systems, composed of histidine kinases and response regulators, enable bacteria to sense, respond, and adapt to changes in their internal and external conditions. The importance of these signaling systems is reflected in their widespread distribution and prevalence in the bacterial ki...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(07)23026-5
更新日期：2007-01-01 00:00:00
abstract：:Fragment-based drug discovery has emerged over the past 15 years as an effective lead discovery paradigm that is complementary to traditional high-throughput screening. The starting point for fragment-based drug discovery is the identification of low-molecular weight, typically low-affinity compounds that bind to a ta...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-381274-2.00006-6
更新日期：2011-01-01 00:00:00
abstract：:Apical caspases 8, 9, and 10 are only active as dimers. These dimers are unstable, and to characterize their activity they need to be maintained in vitro in a dimeric state. We provide updated methods for those looking to characterize various aspects of caspase function. We describe full methods for those looking to a...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-417158-9.00007-8
更新日期：2014-01-01 00:00:00
abstract：:Autophagy is an extremely dynamic process that mediates the rapid degradation of intracellular components in response to different stress conditions. The autophagic response is executed by specific protein complexes, whose function is regulated by posttranslational modifications and interactions with positive and nega...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2016.09.069
更新日期：2017-01-01 00:00:00
abstract：:Microfluidic devices have very broad applications in biological assays from simple chemotaxis assays to much more complicated 3D bioreactors. In this chapter, we describe the design and methods for performing chemotaxis assays using simple microfluidic chemotaxis chambers. With these devices, using real-time video mic...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2015.09.023
更新日期：2016-01-01 00:00:00
abstract：:G-protein-coupled receptors (GPCRs) form a superfamily of membrane proteins that play a crucial role in mediating physiological processes as well as pathogenesis of many critical diseases. They are one of the most successful drug targets, accounting for more than 30% of prescription drugs on the market today. Three-di...
journal_title：Methods in enzymology
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doi：10.1016/S0076-6879(09)05213-6
更新日期：2009-01-01 00:00:00
abstract：:Membrane proteins (MPs) are insoluble in aqueous media as a result of incompatibility between the hydrophilic property of the solvent molecules and the hydrophobic nature of MP surfaces, normally associated with lipid membranes. Amphipathic compounds are necessary for extraction of these macromolecules from the native...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2014.12.021
更新日期：2015-01-01 00:00:00
abstract：:Intracellular calcium (Ca2+) is involved in the regulation of a variety of biological functions in cancer cells, including growth inhibition, tumor invasiveness, and drug resistance. To gain insight into the possible role played by Ca2+ in the development of drug resistance in breast cancer, we performed a comparative...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(06)14018-5
更新日期：2006-01-01 00:00:00
abstract：:Concomitant with the rapid development in biomedical knowledge, including the methods of molecular biology and proteomics, and the manufacture of ever more precise optical instruments, powerful lasers, and sophisticated microcomputing hardware and software, laser microdissection systems have emerged which are now ente...
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doi：10.1016/s0076-6879(02)56935-4
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abstract：:Histone variants and posttranslational modifications (PTMs) are essential for epigenetic regulation of transcriptional expression. Single and/or combinatorial PTMs of histones play important roles in development and disease formation. Mass spectrometry (MS) has been a powerful tool to study histone variants and PTMs, ...
journal_title：Methods in enzymology
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doi：10.1016/B978-0-12-391940-3.00001-9
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abstract：:CRISPR-Cas systems protect prokaryotic cells from invading phages and plasmids by recognizing and cleaving foreign nucleic acid sequences specified by CRISPR RNA spacer sequences. Several CRISPR-Cas systems have been widely used as tool for genetic engineering. In DNA-targeting CRISPR-Cas nucleoprotein effector comple...
journal_title：Methods in enzymology
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abstract：:This article describes the methods and techniques used to produce mutagenized mice to conduct high-throughput forward genetic screens for circadian rhythm mutants in the mouse. In particular, we outline methods to safely prepare and administer the chemical mutagen N-nitroso-N-ethylurea (ENU) to mice. We also discuss t...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)93007-3
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abstract：:Perhaps the most important contribution of FLN is that it provides an experimental approach to relate physical changes in the protein to predicted dynamical behavior. It is clear that the sample is inhomogeneously broadened in a continuous manner, consistent with the damped motion of proteins. At the same time configu...
journal_title：Methods in enzymology
pub_type： 杂志文章,评审
doi：10.1016/s0076-6879(97)78007-8
更新日期：1997-01-01 00:00:00
abstract：:Polyacrylamide gel electrophoresis under native conditions (native PAGE) is a well-established and versatile method for probing nucleic acid conformation and nucleic acid-protein interactions. Native PAGE has been used to measure RNA folding equilibria and kinetics under a wide variety of conditions. Advantages of thi...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(09)69009-1
更新日期：2009-01-01 00:00:00
abstract：:The phenomenon of chemical or conformational exchange in NMR spectroscopy has enabled detailed characterization of time-dependent aspects of biomolecular function, including folding, molecular recognition, allostery, and catalysis, on timescales from microsecond to second. Importantly, NMR methods based on a variety o...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/bs.mie.2018.09.028
更新日期：2019-01-01 00:00:00
abstract：:The Ras-regulatory network is a loosely defined composition of numerous Ras family members and effector pathways that couple to critical cell-regulatory processes. Investigators are increasingly turning to RNAi-mediated inhibition of gene expression as an effective tool to help generate authentic portraits of Ras prot...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/S0076-6879(05)07022-9
更新日期：2006-01-01 00:00:00
abstract：:Physical, chemical, and biological processes commonly discriminate among stable isotopes. Therefore, the stable isotope compositions of biomass, growth substrates, and products often carry the isotopic fingerprints of the processes that shape them. Therefore, measuring isotope fractionation by enzymes and cultures of ...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-385112-3.00014-7
更新日期：2011-01-01 00:00:00
abstract：:Many methods have been developed for the cloning of PCR products. These methods include blunt-end cloning, TA cloning, and using restriction sites incorporated into the PCR primers. The restrictionless cloning technique allows efficient directional cloning of PCR products into any cloning site within a vector regardle...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/B978-0-12-418687-3.00009-4
更新日期：2013-01-01 00:00:00
abstract：:The identification and isolation of the soluble catalytic domains of adenylyl cyclase have provided investigators with useful reagents for the study of these enzymes. They have permitted detailed mechanistic investigation of the actions of forskolin, Gs alpha, and the inhibitory G protein, Gi alpha. They have served a...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/s0076-6879(02)45012-4
更新日期：2002-01-01 00:00:00
abstract：:In mice with lupus nephritis qualitative changes in anti-DNA antibodies occur, such as IgG switch and increased cationic charge, to render these antibodies pathogenic. Pathogenic anti-DNA idiotypes can be encoded by genes of a normal mouse strain such as SWR, where they remain dormant. When the normal mice are crossed...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/0076-6879(88)62094-5
更新日期：1988-01-01 00:00:00
abstract：:Granzyme B (GrB) is the primary molecular mediator of apoptosis by cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. It is a unique mammalian aspartic acid-cleaving serine protease. On T cell receptor activation, GrB is released from the CTL cytoplasmic granules by exocytosis, enters the target cells and, ...
journal_title：Methods in enzymology
pub_type： 杂志文章
doi：10.1016/s0076-6879(00)22013-2
更新日期：2000-01-01 00:00:00
