Design and construction of chimeric linker library with controllable flexibilities for precision protein engineering.

abstract：:Optical imaging of reporter molecules such as firefly luciferase has become a popular method of tracking and visualizing cells in living animals. Many biological processes involve ubiquitin ligases, which target specific proteins for destruction under specific sets of conditions. Importantly, the motifs recognized by ...

journal_title：Methods in enzymology

authors： Zhang GJ,Kaelin WG Jr

更新日期：2005-01-01 00:00:00

abstract：:Staphylococcus aureus is a Gram-positive major human pathogen involved in a wide range of human infectious diseases (from minor skin infections to septicemia, endocarditis or toxic shock syndrome). The treatment of S. aureus infections is very challenging due to the emergence of multiple antibiotic-resistant isolates....

journal_title：Methods in enzymology

authors： Lalaouna D,Desgranges E,Caldelari I,Marzi S

更新日期：2018-01-01 00:00:00

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 bio...

journal_title：Methods in enzymology

authors： Callis PR

更新日期：2011-01-01 00:00:00

abstract：:Endocytosis and autophagy are key vesicular pathways involved in degradation and recycling of cellular material. Both degradative pathways finally fuse with lysosome but are indeed interconnected at several levels. In particular, the fusion between endosomes and autophagosomes can generate intermediate vesicles named ...

journal_title：Methods in enzymology

authors： Manil-Ségalen M,Culetto E,Legouis R,Lefebvre C

更新日期：2014-01-01 00:00:00

abstract：:This chapter presents the application of capillary electrophoresis coupled to electrospray mass spectrometry (CE-ES-MS) for the analysis of complex bacterial lipopolysaccharides (LPS) from pathogenic strains of Haemophilus influenzae and Neisseria meningitidis. A discussion is included of the development of electropho...

journal_title：Methods in enzymology

authors： Li J,Martin A,Cox AD,Moxon ER,Richards JC,Thibault P

更新日期：2005-01-01 00:00:00

abstract：:Dimethylsulfoniopropionate (DMSP) demethylase is a tetrahydrofolate-dependent enzyme that initiates the DMSP demethylation pathway in marine bacteria. This enzyme is important for understanding of organic sulfur flux from the oceans because it directs the sulfur from DMSP away from dimethylsulfide. This enzyme has bee...

journal_title：Methods in enzymology

authors： Reisch CR

更新日期：2018-01-01 00:00:00

abstract：:The field of therapeutic application of the A₃ adenosine receptor (A₃AR) antagonists represents a rapidly growing and intense area of research in the adenosine field. Even if there are currently no A₃AR antagonists in clinical phases, in light of the plethora of biological effects attributed to A₃ARs, substantial effo...

journal_title：Methods in enzymology

authors： Paoletta S,Federico S,Spalluto G,Moro S

更新日期：2010-01-01 00:00:00

abstract：:Measurement of catalysis by MnSOD using direct observation of the UV absorbance of superoxide allows determination of steady-state catalytic constants. Stabilizing superoxide in aprotic solvents such as dimethyl sulfoxide permits the use of stopped-flow spectrophotometry, although significant information is lost in th...

journal_title：Methods in enzymology

authors： Silverman DN,Nick HS

更新日期：2002-01-01 00:00:00

abstract：:Cryptococcus neoformans is a yeastlike fungus that causes a lethal meningoencephalitis in a broad spectrum of immunocompromised patients and has become the most common cause of meningitis due to AIDS-related infections in Africa. Key to the development of new agents to control and prevent this infection is the identif...

journal_title：Methods in enzymology

authors： Hu G,Gibbons J,Williamson PR

更新日期：2008-01-01 00:00:00

abstract：:High content, phenotypic screens offer a powerful approach to systems biology at the cellular level. The approach employs cells carrying fluorescently labeled molecules or organelles in 384- or 1536-well microplates, and an automated confocal screening microscope for capturing images from each well. Although some spec...

journal_title：Methods in enzymology

authors： Kepiro M,Varkuti BH,Davis RL

更新日期：2018-01-01 00:00:00

abstract：:Electron cryo-microscopy (cryoEM) has advanced dramatically to become a viable tool for high-resolution structural biology research. The ultimate outcome of a cryoEM study is an atomic model of a macromolecule or its complex with interacting partners. This chapter describes a variety of algorithms and software to buil...

journal_title：Methods in enzymology

authors： DiMaio F,Chiu W

更新日期：2016-01-01 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

authors： Fay JF,Farrens DL

更新日期：2017-01-01 00:00:00

abstract：:Isothermal Titration Calorimetry (ITC) provides a sensitive and accurate means by which to study the thermodynamics of RNA folding, RNA binding to small molecules, and RNA-protein interactions. The advent of extremely sensitive instrumentation and the increasing availability of ITC in shared facilities have made it in...

journal_title：Methods in enzymology

authors： Feig AL

更新日期：2009-01-01 00:00:00

abstract：:Family 3 carbohydrate-binding modules (CBM3s) are among the most distinctive, diverse, and robust. CBM3s, which are numerous components of both free cellulases and cellulosomes, bind tightly to crystalline cellulose, and thus play a key role in cellulose degradation through their substrate targeting capacity. In addit...

journal_title：Methods in enzymology

authors： Yaniv O,Frolow F,Levy-Assraf M,Lamed R,Bayer EA

更新日期：2012-01-01 00:00:00

abstract：:In actively growing cells, the rate of translation initiation is relatively rapid. As a result, multiple ribosomes simultaneously engaged in translation become spaced along single mRNA molecules. These structures, termed polysomes, can be separated from ribosomal subunits and single ribosomes because they migrate fast...

journal_title：Methods in enzymology

authors： Qin D,Fredrick K

更新日期：2013-01-01 00:00:00

abstract：:Intracellular protein degradation is one of the most precisely regulated processes in living cells. The main component of the degradation machinery is the 20S proteasome present in eukaryotes as well as in prokaryotes. We have developed successful purification protocols for the 20S proteasome in its native state using...

journal_title：Methods in enzymology

authors： Groll M,Huber R

更新日期：2005-01-01 00:00:00

abstract：:Yeast cells in an isogenic population do not all display the same phenotypes. To study such variation within a population of cells, we need to perform measurements on each individual cell instead of measurements that average out the behavior of a cell over the entire population. Here, we provide the basic concepts and...

journal_title：Methods in enzymology

authors： Youk H,Raj A,van Oudenaarden A

更新日期：2010-01-01 00:00:00

abstract：:A genetic interaction occurs when the combination of two mutations leads to an unexpected phenotype. Screens for synthetic genetic interactions have been used extensively to identify genes whose products are functionally related. In particular, synthetic lethal genetic interactions often identify genes that buffer one...

journal_title：Methods in enzymology

authors： Baryshnikova A,Costanzo M,Dixon S,Vizeacoumar FJ,Myers CL,Andrews B,Boone C

更新日期：2010-01-01 00:00:00

abstract：:Specific recognition of mRNA 5' cap by eukaryotic initiation factor eIF4E is a rate-limiting step in the translation initiation. Structural determination of the eIF4E-cap complexes, as well as complexes of eIF4E with other proteins regulating its activity, requires complementary experiments that allow for energetic an...

journal_title：Methods in enzymology

authors： Niedzwiecka A,Stepinski J,Antosiewicz JM,Darzynkiewicz E,Stolarski R

更新日期：2007-01-01 00:00:00

abstract：:Riboswitches present a ubiquitous genetic regulatory mechanism for prokaryotes and have been found in HIV1, fungi, plants, and even H. sapiens. We present an overview of approaches to predict riboswitch aptamers and, more generally, RNA conformational switches. ...

journal_title：Methods in enzymology

authors： Clote P

更新日期：2015-01-01 00:00:00

abstract：:In vivo genome manipulation through site-directed mutagenesis and chromosome rearrangements has been hindered by the difficulty in achieving high frequencies of targeting and the intensive labor required to create altered genomes that do not contain any heterologous sequence. Here we describe our approach, referred to...

journal_title：Methods in enzymology

authors： Storici F,Resnick MA

更新日期：2006-01-01 00:00:00

abstract：:Approximately one-third of all newly translated polypeptides interact with the endoplasmic reticulum (ER), an event that is essential to target these nascent proteins to distinct compartments within the cell or to the extracellular milieu. Thus, the ER houses molecular chaperones that augment the folding of this diver...

journal_title：Methods in enzymology

authors： Brodsky JL

更新日期：2010-01-01 00:00:00

abstract：:Glycosylation is a complex form of protein modification occurring in the secretory pathway. The addition of N- and O-glycans affects intracellular processes like the folding and trafficking of most glycoproteins. To better understand the impact of glycosylation in protein folding and maturation, parameters like glycos...

journal_title：Methods in enzymology

authors： Hülsmeier AJ,Welti M,Hennet T

更新日期：2011-01-01 00:00:00

abstract：:Liquid-liquid phase separation (LLPS) has been known to drive formation of biomolecular compartments, which can encapsulate RNA and proteins among other cosolutes. Such compartments, which lack a lipid membrane, have been implicated in origins of life scenarios as they can easily uptake and concentrate biomolecules, s...

journal_title：Methods in enzymology

authors： Poudyal RR,Meyer MO,Bevilacqua PC

更新日期：2021-01-01 00:00:00

abstract：:Cells belonging to the monocyte/macrophage lineage are in general highly resistant to peroxynitrite. Resistance is not dependent on the scavenging of peroxynitrite itself, or of other secondary reactive species, but is rather associated with the prompt activation of a survival signaling leading to the prevention of to...

journal_title：Methods in enzymology

authors： Cantoni O,Tommasini I,Cerioni L

更新日期：2008-01-01 00:00:00

abstract：:Constitutively activating mutations of the TSHR are the major cause for nonautoimmune hyperthyroidism, which is based on ligand independent, permanent receptor activation. Several reports have highlighted the difficulties to determine whether a TSHR mutation is constitutively active or not especially for borderline ca...

journal_title：Methods in enzymology

authors： Mueller S,Jaeschke H,Paschke R

更新日期：2010-01-01 00:00:00

abstract：:Technical innovations in structural probing have drastically advanced the field of RNA structure analysis. These advances have led to parallel approaches developed in separate labs for analyzing RNA structure and dynamics. With the wealth of methodologies available, it can be difficult to determine which is best suite...

journal_title：Methods in enzymology

authors： Adams RL,Huston NC,Tavares RCA,Pyle AM

更新日期：2019-01-01 00:00:00

abstract：:A fundamental function of the vasculature is to deliver oxygen to tissues and organs. The cells that make up the vasculature also require oxygen, and are acted upon by oxygen in direct and indirect ways that can have significant effects on acute and chronic vascular function and morphology. The role that oxygen, or it...

journal_title：Methods in enzymology

authors： Huang Y,Giordano FJ

更新日期：2008-01-01 00:00:00

abstract：:Extracellular signals dictate the biological processes of neural stem cells (NSCs) both in vivo and in vitro. The intracellular response elicited by these signals is dependent on the context in which the signal is received, which in turn is decided by previous and concurrent signals impinging on the cell. A synthesis ...

journal_title：Methods in enzymology

authors： Rajan P,Snyder E

更新日期：2006-01-01 00:00:00

abstract：:Galectins are a family of animal lectins with affinity for beta-galactosides. By using recombinant proteins, a number of galectins have been shown to interact with cell-surface and extracellular matrix glycoconjugates through lectin-carbohydrate interactions. Through this action, they can affect a variety of cellular ...

journal_title：Methods in enzymology

authors： Hsu DK,Yang RY,Liu FT

更新日期：2006-01-01 00:00:00