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Forster resonance energy transfer

Ting Cheng, Richard O'Rorke, Raphael Francois Ortiz, Tay Yee Yan, Eva Hemmer, Fiorenzo Vetrone, Robert S Marks, Terry W J Steele
Near-infrared (NIR) triggered chemical delivery allows on-demand release with the advantage of external tissue stimulation. Bioresorbable polyester poly-L-lactic acid (PLLA) was compounded with photoadditives of neat zinc oxide (ZnO) nanoparticles and 980 → 365 nm LiYF4:Tm(3+), Yb(3+) upconverting nanoparticles (UCNP). Subsequently, neat ZnO and UCNP blended PLLA films of sub-50 μm thickness were knife casted with a hydrophobic small molecule drug mimic, fluorescein diacetate. The PLLA films displayed a 500 times increase in fluorescein diacetate release from the 50 mW NIR irradiated PLLA/photoadditive film compared to non-irradiated PLLA control films...
March 15, 2017: Acta Biomaterialia
Andreas Schlundt, Jan-Niklas Tants, Michael Sattler
Recent advances in RNA sequencing technologies have greatly expanded our knowledge of the RNA landscape in cells, often with spatiotemporal resolution. These techniques identified many new (often non-coding) RNA molecules. Large-scale studies have also discovered novel RNA binding proteins (RBPs), which exhibit single or multiple RNA binding domains (RBDs) for recognition of specific sequence or structured motifs in RNA. Starting from these large-scale approaches it is crucial to unravel the molecular principles of protein-RNA recognition in ribonucleoprotein complexes (RNPs) to understand the underlying mechanisms of gene regulation...
March 15, 2017: Methods: a Companion to Methods in Enzymology
Marijonas Tutkus, Tomas Marciulionis, Giedrius Sasnauskas, Danielis Rutkauskas
The single-molecule Förster resonance energy transfer (FRET) is a powerful tool to study interactions and conformational changes of biological molecules in the distance range from a few to 10 nm. In this study, we demonstrate a method to augment this range with longer distances. The method is based on the intensity changes of a tethered fluorophore, diffusing in the exponentially decaying evanescent excitation field. In combination with FRET it allowed us to reveal and characterize the dynamics of what had been inaccessible conformations of the DNA-protein complex...
March 14, 2017: Biophysical Journal
Rolf Lood, Henrik Molina, Vincent A Fischetti
The necessity of identifying novel methods to combat infections caused by antibiotic resistant bacteria is increasing each year. Recent advancements in the development of peptidoglycan hydrolases (e.g. lysins) from bacterial viruses (bacteriophages) have revealed the efficiency of this class of enzymes in treating serious bacterial infections. Though promising results have been obtained regarding the lethal action of lysin on bacterial pathogens both in vitro and in vivo, an often-overlooked factor in these studies is precisely identifying their peptidoglycan cleavage site...
2017: PloS One
Falk Schneider, Daja Ruhlandt, Ingo Gregor, Jörg Enderlein, Alexey I Chizhik
Precise knowledge of the quantum yield is important for many fluorescence-spectroscopic techniques, for example, for Förster resonance energy transfer. However, to measure it for emitters in a complex environment and at low concentrations is far from being trivial. Using a plasmonic nanocavity, we measure the absolute quantum yield value of lipid-conjugated dyes incorporated into a supported lipid bilayer. We show that for both hydrophobic and hydrophilic molecules the quantum yield of dyes inside the lipid bilayer strongly differs from its value in aqueous solution...
March 20, 2017: Journal of Physical Chemistry Letters
Zhi Zhou Li, Feng Liang, Ming Peng Zhuo, Ying Li Shi, Xue Dong Wang, Liang Sheng Liao
Organic semiconductor micro-/nanocrystals with regular shapes have been demonstrated for many applications, such as organic field-effect transistors, organic waveguide devices, organic solid-state lasers, and therefore are inherently ideal building blocks for the key circuits in the next generation of miniaturized optoelectronics. In the study, blue-emissive organic molecules of 1,4-bis(2-methylstyryl)benzene (o-MSB) can assemble into rectangular microcrystals at a large scale via the room-temperature solution-exchange method...
March 10, 2017: Small
Yuki Ohmuro-Matsuyama, Hiroshi Ueda
We recently developed a protein-protein interaction assay, FlimPIA (Firefly luminescent intermediate-based Protein-protein Interaction Assay) based on the catalytic mechanism of firefly luciferase (Fluc) that can be divided into two half-reactions: the adenylation step and the oxidative luminescent steps. We engineered two mutant Fluc enzymes named "Donor" and "Acceptor" where the oxidative luminescent activity of the Donor is almost eliminated and the adenylation activity of the Acceptor is suppressed. When the Donor and the Acceptor are each fused to one of two interacting partners, and put together to interact, the Donor and the Acceptor come sufficiently close such that the Acceptor can react with the luciferyl-adenylate intermediate (LH2-AMP) produced by the Donor, and thus emit luminescence...
2017: Methods in Molecular Biology
Joshua A Mitchell, William H Zhang, Michel K Herde, Christian Henneberger, Harald Janovjak, Megan L O'Mara, Colin J Jackson
Biosensors that exploit Förster resonance energy transfer (FRET) can be used to visualize biological and physiological processes and are capable of providing detailed information in both spatial and temporal dimensions. In a FRET-based biosensor, substrate binding is associated with a change in the relative positions of two fluorophores, leading to a change in FRET efficiency that may be observed in the fluorescence spectrum. As a result, their design requires a ligand-binding protein that exhibits a conformational change upon binding...
2017: Methods in Molecular Biology
Ben E Clifton, Jason H Whitfield, Inmaculada Sanchez-Romero, Michel K Herde, Christian Henneberger, Harald Janovjak, Colin J Jackson
Small molecule biosensors based on Förster resonance energy transfer (FRET) enable small molecule signaling to be monitored with high spatial and temporal resolution in complex cellular environments. FRET sensors can be constructed by fusing a pair of fluorescent proteins to a suitable recognition domain, such as a member of the solute-binding protein (SBP) superfamily. However, naturally occurring SBPs may be unsuitable for incorporation into FRET sensors due to their low thermostability, which may preclude imaging under physiological conditions, or because the positions of their N- and C-termini may be suboptimal for fusion of fluorescent proteins, which may limit the dynamic range of the resulting sensors...
2017: Methods in Molecular Biology
Jeung-Hoi Ha, Stewart N Loh
Alternate frame folding (AFF) and protein/fragment exchange (FREX) are related technologies for engineering allosteric conformational changes into proteins that have no pre-existing allosteric properties. One of their chief purposes is to turn an ordinary protein into a biomolecular switch capable of transforming an input event into an optical or functional readout. Here, we present a guide for converting an arbitrary binding protein into a fluorescent biosensor with Förster resonance energy transfer output...
2017: Methods in Molecular Biology
Bo Bai, Xiaoyu Chen, Rumin Zhang, Xin Wang, Yunlu Jiang, Dandan Li, Zhengwen Wang, Jing Chen
The orexin (OX1R) and cholecystokinin A (CCK1R) receptors play opposing roles in the migration of the human colon cancer cell line HT-29, and may be involved in the pathogenesis and pathophysiology of cancer cell invasion and metastasis. OX1R and CCK1R belong to family A of the G-protein-coupled receptors (GPCRs), but the detailed mechanisms underlying their functions in solid tumor development remain unclear. In this study, we investigated whether these two receptors heterodimerize, and the results revealed novel signal transduction mechanisms...
March 10, 2017: Biochimica et Biophysica Acta
Yuanqing Ma, Yui Yamamoto, Philip R Nicovich, Jesse Goyette, Jérémie Rossy, J Justin Gooding, Katharina Gaus
Membrane charge has a critical role in protein trafficking and signaling. However, quantification of the effective electrostatic potential of cellular membranes has remained challenging. We developed a fluorescence membrane charge sensor (MCS) that reports changes in the membrane charge of live cells via Förster resonance energy transfer (FRET). MCS is permanently attached to the inner leaflet of the plasma membrane and shows a linear, reversible and fast response to changes of the electrostatic potential...
March 13, 2017: Nature Biotechnology
Mengqun Yu, Hong Wang, Fei Fu, Linyao Li, Jing Li, Gan Li, Yang Song, Mark T Swihart, Erqun Song
The effective monitoring, identification, and quantification of pathogenic bacteria is essential for addressing serious public health issues. In this study, we present a universal and facile one-step strategy for sensitive and selective detection of pathogenic bacteria using a dual molecular affinity-based Förster (fluorescence) resonance energy transfer (FRET) platform based on the recognition of bacterial cell walls by antibiotic and aptamer molecules, respectively. As a proof of concept, Vancomycin (Van), and a nucleic acid aptamer were employed in a model dual recognition scheme for detecting Staphylococcus aureus (S...
March 13, 2017: Analytical Chemistry
Okhil K Nag, Jawad Naciri, Eunkeu Oh, Christopher M Spillmann, James B Delehanty
The controlled delivery of drug/imaging agents to cells is critical for the development of therapeutics and for the study of cellular signaling processes. Recently, nanoparticles (NPs) have shown significant promise in the development of such delivery systems. Here, a liquid crystal NP (LCNP)-based delivery system has been employed for the controlled delivery of a water-insoluble dye, 3,3'-dioctadecyloxacarbocyanine perchlorate (DiO), from within the NP core to the hydrophobic region of a plasma membrane bilayer...
February 8, 2017: Journal of Visualized Experiments: JoVE
Thilo Dörfler, Tobias Eilert, Carlheinz Röcker, Julia Nagy, Jens Michaelis
Single-molecule Förster Resonance Energy Transfer (smFRET) can be used to obtain structural information on biomolecular complexes in real-time. Thereby, multiple smFRET measurements are used to localize an unknown dye position inside a protein complex by means of trilateration. In order to obtain quantitative information, the Nano-Positioning System (NPS) uses probabilistic data analysis to combine structural information from X-ray crystallography with single-molecule fluorescence data to calculate not only the most probable position but the complete three-dimensional probability distribution, termed posterior, which indicates the experimental uncertainty...
February 9, 2017: Journal of Visualized Experiments: JoVE
Thibaut Deschamps, Quentin Bazot, Derek M Leske, Ruth MacLeod, Dimitri Mompelat, Lionel Tafforeau, Vincent Lotteau, Vincent Maréchal, George S Baillie, Henri Gruffat, Joanna B Wilson, Evelyne Manet
The Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is a sequence-specific DNA-binding protein that plays an essential role in viral episome replication and segregation, by recruiting the cellular complex of DNA replication onto the origin (oriP) and by tethering the viral DNA onto the mitotic chromosomes. Whereas the mechanisms of viral DNA replication are well documented, those involved in tethering EBNA1 to the cellular chromatin are far from being understood. Here, we have identified regulator of chromosome condensation 1 (RCC1) as a novel cellular partner for EBNA1...
February 2017: Journal of General Virology
Philipp Heissig, Waldemar Schrimpf, Philipp Hadwiger, Ernst Wagner, Don C Lamb
Short single-stranded oligonucleotides represent a class of promising therapeutics with diverse application areas. Antisense oligonucleotides, for example, can interfere with various processes involved in mRNA processing through complementary base pairing. Also RNA interference can be regulated by antagomirs, single-stranded siRNA and single-stranded microRNA mimics. The increased susceptibility to nucleolytic degradation of unpaired RNAs can be counteracted by chemical modification of the sugar phosphate backbone...
2017: PloS One
Tomoki Yoshida, Soaad Alfaqaan, Norio Sasaoka, Hiromi Imamura
Genetically encoded biosensors utilizing the Förster resonance energy transfer (FRET) are powerful tools for live cell imaging of various cellular processes. Our group has previously developed a series of FRET-based biosensors, named "ATeam," for visualization of ATP levels inside a single living cell. ATeam not only provides a window of insight into a single cell but also allows for visualization of ATP levels in mitochondrial matrix of a single living cell. This novel tool is able to monitor alterations in cellular ATP in response to various treatments in real time...
2017: Methods in Molecular Biology
Emilie Roger, Jean-Christophe Gimel, Conor Bensley, Andrey S Klymchenko, Jean-Pierre Benoit
Lipid nanocapsules (LNCs) have demonstrated great potential for the oral delivery of drugs having very limited oral bioavailability (BCS class II, III and IV molecules). It has been shown previously that orally-administered LNCs can permeate through mucus, increase drug absorption by the epithelial tissue, and finally, increase drug bioavailability. However, even if transport mechanisms through mucus and the intestinal barrier have already been clarified, the preservation of particle integrity is still not known...
March 6, 2017: Journal of Controlled Release: Official Journal of the Controlled Release Society
Etienne Gagnon, Audrey Connolly, Jessica Dobbins, Kai W Wucherpfennig
Over the last decade, advancements in the time and space resolution of microscopy technologies have enabled dissection of the molecular events involved in T cell Immunological Synapse (IS) formation. Using a combination of Förster Resonance Energy Transfer (FRET) and Fluorescence Lifetime Imagining Microscopy (FLIM), we have demonstrated dynamic plasma membrane binding by cytoplasmic domains of T cell receptor (TCR)-associated CD3 chains and other T cell transmembrane receptors. We have developed methods for imaging such membrane binding both at steady state and during receptor triggering at the IS...
2017: Methods in Molecular Biology
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