Abstract:
:Model building studies may be used to supplement structurally low resolution experimental data with detailed three-dimensional hypothetical atomic models. Because of the strong relation between structure and function in biological molecules such models may give a consistent, integral view of a wealth of experimental data. In most cases such models will predict the outcome of certain experiments. The outcome of these experiments will often either confirm the model may be used for further refinement or even demand a major revision of the model. Coordinates obtained from X-ray fiber diffraction data or in special cases single-crystal data may provide the elements for DNA or RNA model building. Local and nonlocal optimization may be used to refine these structures and to evaluate their statistical significance as estimated by a chosen force field. Appreciable progress using nonlocal optimization procedures can only be expected if the dimensionality of the problem can be reduced sufficiently to the relevant degrees of freedom. Taking advantage of structural symmetries may critically improve the convergence while refining the target molecule or its building blocks. Monte Carlo and molecular dynamics methods allow one to calculate averaged quantities. In addition, molecular dynamics provides time evolutions of certain averages. During the simulation of certain physical properties of molecules a huge amount of data will be generated. They will provide many answers, but these answers may not always apply to the original question. So what type of questions will be reliably answered by a force field? Relatively safe answers concern the local geometry of the molecules. If a conformation leads to strong distortions of bond distances or angles or to close van der Waals contacts, this conformation can safely be rejected. Optimizing such unfavorable structures energetically may lead to structures showing how to avoid such distortions. More difficult are energetical questions: which of two conformers is more stable, or what is the free energy of the substrate in the active site? One cannot always be sure that the force field provides the correct answer. Therefore, one should pose only those questions which can be checked experimentally. Because of the many possible answers, the experiment may benefit by starting with a choice proposed by the simulation. The application of this procedure to curved DNA and the DNA four-way junction was successful.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
von Kitzing Edoi
10.1016/0076-6879(92)11024-dsubject
Has Abstractpub_date
1992-01-01 00:00:00pages
449-67eissn
0076-6879issn
1557-7988journal_volume
211pub_type
杂志文章,评审abstract::Portable, nanoparticle (NP)-enhanced enzyme sensors have emerged as powerful devices for qualitative and quantitative analysis of a variety of analytes for biomedicine, environmental applications, and pharmaceutical fields. This chapter describes a method for the fabrication of a portable, paper-based, inexpensive, ro...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2016.03.006
更新日期:2016-01-01 00:00:00
abstract::Microsatellite markers are quickly becoming the molecular marker of choice for studies of parentage and kinship in animals. In this chapter, we review methods and give protocols for screening potential microsatellite markers, as well as protocols for genotyping individuals with useful markers once they have been ident...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(05)95014-3
更新日期:2005-01-01 00:00:00
abstract::Epitope tagged alpha 2-AR subtypes have been used to address a variety of cell biological questions, and the strategies used are readily applicable to all GPCR as well as other cell surface proteins. We have provided detailed protocols for successful utilization of the epitope-tagged receptor in the studies of protein...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(02)43156-4
更新日期:2002-01-01 00:00:00
abstract::Protein (poly-)ubiquitination is a posttranslational modification that plays a key role in almost all cellular processes. It involves the installment of either single ubiquitin (Ub) moieties or one of eight different polyUb linkage types, each giving a distinct cellular outcome. Deubiquitinating enzymes (DUBs) reverse...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2018.12.037
更新日期:2019-01-01 00:00:00
abstract::The study of hormone secretion by anterior pituitary cells is complicated by the presence of multiple cell types. For unambiguous interpretation of data it is necessary to identify the cells from which measurements are made. We have described a reliable experimental approach involving the identification of cultured ce...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/0076-6879(89)68011-1
更新日期:1989-01-01 00:00:00
abstract::The reverse dot-blot method is a simple and rapid diagnostic procedure that allows screening of sample for a variety of mutations/polymorphisms in a single hybridization reaction. Several methods of immobilizing the oligonucleotide probes are discussed. The reverse dot-blot method has several unique properties that ar...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/0076-6879(93)18029-c
更新日期:1993-01-01 00:00:00
abstract::Cholestatic syndromes are inborn or acquired disorders of bile formation. In recent years, several inherited cholestatic syndromes were characterized at the molecular level: progressive familial intrahepatic cholestasis (PFIC) and benign recurrent intrahepatic cholestasis (BRIC). Both PFIC and BRIC were divided phenot...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(05)00031-5
更新日期:2005-01-01 00:00:00
abstract::Death receptors are a subgroup of the tumor necrosis factor receptor superfamily (TNFRSF) and mediate activation of what is widely known as the "extrinsic" apoptosis pathway. TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) is one of the most recent death receptor ligands identified. The TRAIL receptor ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(08)01618-2
更新日期:2008-01-01 00:00:00
abstract::Nedd8 is a ubiquitin-like molecule that is highly conserved in eukaryotes. Similar to ubiquitin, Nedd8 attaches to target proteins through an enzymatic cascade composed of Nedd8-specific E1 (activating)- and E2 (conjugating)-enzymes. The E1 for Nedd8 is a heterodimer of APP-BP1 and Uba3, while the E2 is Ubc12. The mos...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(05)98007-5
更新日期:2005-01-01 00:00:00
abstract::β-arrestins (βarrs) are multifunctional proteins that interact with activated and phosphorylated G protein-coupled receptors (GPCRs) to regulate their signaling and trafficking. Understanding the intricate details of GPCR-βarr interaction continues to be a key research area in the field of GPCR biology. Bimane fluores...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2019.11.009
更新日期:2020-01-01 00:00:00
abstract::Since the onset of the genomic era more than 1000 bacterial genomes have been sequenced and several fold more are expected to be completed in the near future. These genome sequences supply a wealth of information that can be exploited by statistical methods to gain significant insights into cellular processes. In Volu...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)71002-8
更新日期:2010-01-01 00:00:00
abstract::Mathematical computer-assisted simulations of episodic hormone, substrate, or metabolite release can be accomplished using explicit algebra and high-speed microprocessors. Such simulations are useful to describe the behavior of single or multiple pulse generators, as well as the expected output of these systems. Simul...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(94)40056-3
更新日期:1994-01-01 00:00:00
abstract::Proteins change their charge state through protonation and redox reactions as well as through binding charged ligands. The free energy of these reactions is dominated by solvation and electrostatic energies and modulated by protein conformational relaxation in response to the ionization state changes. Although computa...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/bs.mie.2016.05.052
更新日期:2016-01-01 00:00:00
abstract::After cell lysis, the most often used second step in a protein purification procedure is some sort of a rapid, bulk precipitation step. This is commonly accomplished by altering the solvent conditions and taking advantage of the changes in solubility of your protein of interest relative to those of many of the other p...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(09)63020-2
更新日期:2009-01-01 00:00:00
abstract::Anomalous small angle X-ray scattering (ASAXS) exploits contrast variation methods to highlight the scattering from one elemental component in a multielement sample, such as one ion species in an ion-DNA system. The ASAXS method has been applied to measure ions condensed around short nucleic acid duplexes. This chapte...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(09)69019-4
更新日期:2009-01-01 00:00:00
abstract::Small GTPases of the Ras and Rho families are among the most studied signaling proteins and represent promising therapeutic targets for human neoplastic disease. Despite the high level of interest in these proteins, direct analysis of most aspects of Ras protein biology in living cells has not been possible, because m...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(05)07032-1
更新日期:2006-01-01 00:00:00
abstract::ADP-ribosylation factor domain protein 1 (ARD1) is a multifunctional protein that belongs to the family of 20-kDa ARF proteins. The ARD1 gene encodes a 64-kDa protein with a structure comprising an 18-kDa ADP-ribosylation factor (ARF) domain at the C-terminus (amino acids 403-574), and a 46-kDa N-terminal domain (amin...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(05)04019-X
更新日期:2005-01-01 00:00:00
abstract::Accurate diagnosis of attentional disorders such as attention-deficit hyperactivity disorder (ADHD) is imperative because there are multiple negative psychosocial sequelae related to undiagnosed and untreated ADHD. Early and accurate detection can lead to effective intervention and prevention of negative sequelae. Unf...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(09)67014-2
更新日期:2009-01-01 00:00:00
abstract::The methods described here concern the use specialized glassware to achieve accurate sub-micromolar to low-millimolar dioxygen concentrations specifically for the study of flavoproteins. Since the early 1960s, these methods have been passed through the lineage of flavin enzymologists almost exclusively as an oral trad...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2019.03.005
更新日期:2019-01-01 00:00:00
abstract::Recent studies using sulfotransferase-deficient mice have revealed various physiological functions of sulfated glycans. Studies using gene-targeted mice deficient in both N-acetylglucosamine-6-O-sulfotransferase (GlcNAc6ST)-1 and GlcNAc6ST-2 showed that these sulfotransferases play critical roles in lymphocyte homing....
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)79014-5
更新日期:2010-01-01 00:00:00
abstract::Hormone, hormone-like substances, and neuroendocrine peptides are natural constituents of milk, and they may have an important physiological and pharmacological role in neonate development. The concentration of these peptides in milk greatly exceeds those in serum and implies an active concentration mechanism in the m...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/0076-6879(89)68038-x
更新日期:1989-01-01 00:00:00
abstract::The Escherichia coli fumarate and nitrate reductase (FNR) regulator protein is an important transcriptional regulator that controls the expression of a large regulon of more than 100 genes in response to changes in oxygen availability. FNR is active when it acquires a [4Fe-4S](2+) cluster under anaerobic conditions. T...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(07)37011-0
更新日期:2008-01-01 00:00:00
abstract::Nonsense-mediated mRNA decay (NMD) is activated by exon-junction complexes (EJCs) that are located downstream of the termination codon of the substrate mRNAs. This situation can be imitated by tethering components of the EJC to the 3' untranslated region (3' UTR) of a reporter mRNA. Here we describe the detailed use o...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(08)02623-2
更新日期:2008-01-01 00:00:00
abstract::Site-directed mutagenesis is a PCR-based method to mutate specified nucleotides of a sequence within a plasmid vector. This technique allows one to study the relative importance of a particular amino acid for protein structure and function. Typical mutations are designed to disrupt or map protein-protein interactions,...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/B978-0-12-418687-3.00019-7
更新日期:2013-01-01 00:00:00
abstract::Proteins are fragile molecules that often require great care during purification to ensure that they remain intact and fully active. Nowadays, many proteins are also purified in small amounts under denaturing conditions by various gel electrophoretic techniques, such that inactive proteins are obtained. But even here,...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(09)63010-X
更新日期:2009-01-01 00:00:00
abstract::Graphene oxide (GO) and reduced graphene oxide (rGO) were demonstrated in the past decade as biocompatible carbon-based materials that could be efficiently used in bioelectrochemical systems (BESs). Specifically, for redox enzyme encapsulation in order to improve electron communication between enzymes and electrodes. ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2018.05.008
更新日期:2018-01-01 00:00:00
abstract::Three-dimensional (3D) cryoelectron microscopy reconstruction methods are uniquely able to reveal structures of many important macromolecules and macromolecular complexes. EMDataBank.org, a joint effort of the Protein Databank in Europe (PDBe), the Research Collaboratory for Structural Bioinformatics (RCSB), and the N...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)83004-6
更新日期:2010-01-01 00:00:00
abstract::Epitope tagging has provided a useful experimental strategy with widespread applicability. The ample variety of epitope tag systems that have been put to use to date provide a collection of attributes relevant to virtually any experimental system. As a consequence, epitope tagging will continue to be a valuable tool f...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/s0076-6879(00)27263-7
更新日期:2000-01-01 00:00:00
abstract::The human cannabinoid receptor, CB1, has been difficult to purify in a functional form, hampering structural and biophysical studies. Here, we present our approaches for obtaining pure, detergent solubilized, functional CB1. We also discuss our site-directed fluorescence labeling (SDFL) methods for identifying differe...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2017.06.026
更新日期:2017-01-01 00:00:00
abstract::IF-liposomes are formed by a unique process that involves fusing small liposomes into interdigitated lipid sheets, using either ethanol or hydrostatic pressure. The interdigitation-fusion method requires liposome formulations with lipids that form the L beta I phase. Preparing ethanol-induced IF-liposomes is simple an...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(03)67007-2
更新日期:2003-01-01 00:00:00