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Forster resonance energy transfer

Yavuz S Dagdas, Janice S Chen, Samuel H Sternberg, Jennifer A Doudna, Ahmet Yildiz
The Cas9 endonuclease is widely used for genome engineering applications by programming its single-guide RNA, and ongoing work is aimed at improving the accuracy and efficiency of DNA targeting. DNA cleavage of Cas9 is controlled by the conformational state of the HNH nuclease domain, but the mechanism that governs HNH activation at on-target DNA while reducing cleavage activity at off-target sites remains poorly understood. Using single-molecule Förster resonance energy transfer, we identified an intermediate state of Streptococcus pyogenes Cas9, representing a conformational checkpoint between DNA binding and cleavage...
August 2017: Science Advances
David A Murison, Rebecca C Timson, Bilyana Koleva, Michael Ordazzo, Penny J Beuning
The Escherichia coli SOS response, an induced DNA damage response pathway, confers survival on bacterial cells by providing accurate repair mechanisms as well as the potentially mutagenic pathway translesion synthesis (TLS). The umuD gene products are upregulated after DNA damage and play roles in both non-mutagenic and mutagenic aspects of the SOS response. Full-length UmuD is expressed as a homodimer of 139-amino-acid subunits, which eventually cleaves its N-terminal 24 amino acids to form UmuD'. The cleavage product, UmuD', together with UmuC, form the Y-family polymerase DNA Pol V (UmuD'2C) capable of performing TLS...
August 14, 2017: Biochemistry
Daniel C Sultanov, Nadezhda S Gerasimova, Kseniya S Kudryashova, Natalya V Maluchenko, Elena Y Kotova, Marie-France Langelier, John M Pascal, Mikhail P Kirpichnikov, Alexey V Feofanov, Vasily M Studitsky
DNA accessibility to various protein complexes is essential for various processes in the cell and is affected by nucleosome structure and dynamics. Protein factor PARP-1 (poly(ADP-ribose)polymerase 1) increases the accessibility of DNA in chromatin to repair proteins and transcriptional machinery, but the mechanism and extent of this chromatin reorganization are unknown. Here we report on the effects of PARP-1 on single nucleosomes revealed by spFRET (single-particle Förster Resonance Energy Transfer) microscopy...
2017: AIMS Genetics
Carolina Sanchez, Lena Voith-von-Voithenberg, Lisa Warner, Don C Lamb, Michael Sattler
Fluorescence-based techniques are widely used to study biomolecular conformations, intra- and intermolecular interactions, and conformational dynamics of macromolecules. Especially for fluorescent-based single-molecule experiments, the choice of the fluorophore and labeling position are highly important. Here, we have studied the biophysical and structural effects that are associated with the conjugation of fluorophores to cysteines in the splicing factor U2AF65 by using single pair Förster resonance energy transfer (FRET) and nuclear magnetic resonance (NMR) spectroscopy...
August 10, 2017: Chemistry: a European Journal
Zhen Sheng, Ligang Chen
The concentration of L-cysteine (Cys) and glutathione (GSH) is closely related to the critical risk of various diseases. In our study, a new rapid method for the determination of Cys and GSH in water and urine samples has been developed using a fluorescent probe technique, which was based on crystal violet (CV)-functionalized CdTe quantum dots (QDs). The original QDs emitted fluorescence light, which was turned off upon adding CV. This conjugation of CV and QDs could be attributed to electrostatic interaction between COO(-) of mercaptopropionic acid (MPA) on the surface of QDs and N(+) of CV in aqueous solution...
August 10, 2017: Analytical and Bioanalytical Chemistry
Qi Zhang, Sudipta Lahiri, Tithi Banerjee, Zhongmou Sun, Donald Oliver, Ishita Mukerji
A conserved hairpin-like structure comprised of a signal peptide and early mature region initiates protein transport across the SecY or Sec61α channel in Bacteria or Archaea and Eukarya, respectively. When and how this initiator substrate hairpin forms remains a mystery. Here, we have used the bacterial SecA ATPase motor protein and SecYEG channel complex to address this question. Engineering of a functional miniprotein substrate onto the end of SecA allowed us to efficiently form ternary complexes with SecYEG for spectroscopic studies...
August 10, 2017: Proceedings of the National Academy of Sciences of the United States of America
Stephen E McGowan, Diann M McCoy
Platelet-derived growth factor (PDGF)-A, which only signals through PDGF-receptor-alpha (PDGFRα) is required for secondary alveolar septal formation. Although PDGFRα distinguishes mesenchymal progenitor cells during the saccular stage, PDGFRα-expressing alveolar cells persist through adulthood. PDGF-A sustains proliferation, limits apoptosis, and maintains alpha-smooth muscle actin (αSMA) containing alveolar cells, which congregate at the alveolar entry ring at postnatal day (P)12. PDGFRα-expressing, αSMA-containing, alveolar cells re-distribute in the elongating septum, suggesting that they migrate to the alveolar entry rings where mechanical tension is higher...
August 3, 2017: American Journal of Physiology. Lung Cellular and Molecular Physiology
Dennis K P Ng, Roy Wong, Wing-Ping Fong, Pui-Chi Lo
Doxorubicin (Dox) was conjugated to a zinc(II) phthalocyanine (ZnPc) via an acid-cleavable hydrazone linker. This conjugate, having an azido group, was then anchored to the nanochannels of an alkyne-modified mesoporous silica nanoparticle (MSN) system via copper(I)-catalyzed azide-alkyne cycloaddition reaction. An analogous nanosystem was also prepared by immobilization of a hydrazine-substituted ZnPc to the MSN followed by coupling with Dox for comparison. The release of Dox under acidic conditions was studied in phosphate buffered saline with 0...
August 3, 2017: Chemistry: a European Journal
Handan Acar, Ravand Samaeekia, Matthew R Schnorenberg, Dibyendu K Sasmal, Jun Huang, Matthew V Tirrell, James L LaBelle
Peptides synthesized in the likeness of their native interaction domain(s) are natural choices to target protein-protein interactions (PPIs) due to their fidelity of orthostatic contact points between binding partners. Despite therapeutic promise, intracellular delivery of biofunctional peptides at concentrations necessary for efficacy remains a formidable challenge. Peptide Amphiphiles (PAs) provide a facile method of intracellular delivery and stabilization of bioactive peptides. PAs consisting of biofunctional peptide headgroups linked to hydrophobic alkyl lipid-like tails prevent peptide hydrolysis and proteolysis in circulation and PA monomers are internalized via endocytosis...
August 3, 2017: Bioconjugate Chemistry
Hassan Mansouri-Torshizi, Fatemeh Khosravi, Arezou Ghahghaei, Somaye Shahraki, Sareh Zareian-Jahromi
Two Zn(II) complexes of formula [Zn(bpy)(Gly)]NO3 (I) and [Zn(phen)(Gly)]NO3 (II) (where bpy=2,2'-bipyridine, phen=1,10-phenanthroline and Gly=glycine) were synthesized and characterized by elemental analysis, molar conductance measurements, UV-Vis, FT-IR and (1)H NMR spectra. The interaction ability of these complexes with calf thymus DNA was monitored using spectroscopic methods, including UV-Vis absorption spectroscopy, ethidium bromide displacement, Fourier transform infrared and electrophoretic mobility assay...
August 3, 2017: Journal of Biomolecular Structure & Dynamics
Ying Li, Anyi Wang, Yunfei Bai, Shiping Wang
A novel solid-surface fluorescence biosensor for rapid detection of Sudan I-IV was proposed based on fluorescence resonance energy transfer (FRET). The biosensor was fabricated by immobilizing acriflavine (AY) on the eggshell membrane (ESM) with glutaraldehyde as cross-linking agent. FRET mechanism was demonstrated by using AY and Sudan dyes as donor and acceptor respectively, an efficient energy transfer in the present system was indicated by the sufficient spectral overlap integral (J) and proper Förster critical distance (R0)...
December 15, 2017: Food Chemistry
Lei Yang, Wenxing Gu, Ling Hong, Yang Mi, Feng Liu, Ming Liu, Yufei Yang, Bigyan Sharma, Xinfeng Liu, Hui Huang
Nonradiative Förster resonance energy transfer (FRET) is an important mechanism of organic solar cells, which can improve the exciton migration over a long distance, resulting in improvement of efficiency of solar cells. However, the current observations of FRET are very limited, and the efficiencies are less than 9%. In this study, FRET effect was first observed between two nonfullerene acceptors in ternary solar cells, which improved both the absorption range and exciton harvesting, leading to the dramatic enhancement in the short circuit current and power conversion efficiency...
August 16, 2017: ACS Applied Materials & Interfaces
Suhui Deng, Jianfang Chen, Zhaoshuai Gao, Chunhai Fan, Qiurong Yan, Yuhao Wang
Förster resonance energy transfer (FRET) probes being used to improve the resolution of stimulated emission depletion (STED) microscopy are numerically discussed. Besides the FRET efficiency and the excitation intensity, the fluorescence lifetimes of donor and acceptor are found to be another key parameter for the resolution enhancement. Using samples of FRET pairs with shorter donor lifetime and longer acceptor lifetime enhances the nonlinearity of the donor fluorescence, which leads to an increased resolution...
July 31, 2017: Journal of Microscopy
Mona Mittal, Sameer Sapra
One of the recent developments in enhancing the performance of quantum dot sensitized solar cells (QDSSCs) is to combine QDs with dyes in order to overcome the drawbacks of QDSSCs. However, implementation of this requires a detailed investigation of the interaction between QDs and dye. Here, we have studied the effect of size and surface ligands in the interaction of CdSe nanocrystals (NCs) with Ru N-719 dye. The interaction mechanism is investigated by steady-state and time-resolved photoluminescence spectroscopy, indicating the involvement of apparent static as well as dynamic quenching...
July 31, 2017: Chemphyschem: a European Journal of Chemical Physics and Physical Chemistry
Hideji Murakoshi, Akihiro C E Shibata
Fluorescence lifetime imaging microscopy (FLIM)-based Förster resonance energy transfer (FRET) measurement (FLIM-FRET) is one of the powerful methods for imaging of intracellular protein activities such as protein-protein interactions and conformational changes. Here, using saturation mutagenesis, we developed a dark yellow fluorescent protein named ShadowY that can serve as an acceptor for FLIM-FRET. ShadowY is spectrally similar to the previously reported dark YFP but has a much smaller quantum yield, greater extinction coefficient, and superior folding property...
July 28, 2017: Scientific Reports
Sarah V Faull, Anwen E Brown, Imran Haq, James A Irving
Many serpinopathies, including alpha-1 antitrypsin (A1AT) deficiency, are associated with the formation of unbranched polymer chains of mutant serpins. In vivo, this deficiency is the result of mutations that cause kinetic or thermodynamic destabilization of the molecule. However, polymerization can also be induced in vitro from mutant or wild-type serpins under destabilizing conditions. The characteristics of the resulting polymers are dependent upon induction conditions. Due to their relationship to disease, serpin polymers, mainly those formed from A1AT, have been widely studied...
2017: Methods in Molecular Biology
Qing Wang, Min Pan, Jie Wei, Xiaoqing Liu, Fuan Wang
Methyltransferase (MTase)-catalyzed DNA methylation plays a vital role in the biological epigenetic processes of key diseases and has attracted increasing attention, making the amplified detection of MTase activity of great significance in clinical disease diagnosis and treatment. Herein, we developed an isothermal, enzyme-free, and autonomous strategy for analyzing MTase activity based on concatenated hybridization chain reaction (C-HCR)-mediated Förster resonance energy transfer (FRET). In a typical C-HCR procedure without MTase (Dam), Y-shaped initiator DNA activates upstream HCR-1 to assemble a double-stranded DNA (dsDNA) copolymeric nanowire consisting of multiple tandem DNA trigger units that motivate downstream HCR-2 to successively bring a fluorophore donor/acceptor (FAM/TAMRA) pair into close proximity, leading to the generation of an amplified FRET readout signal...
July 28, 2017: ACS Sensors
Yuchen Long, Yvonne Stahl, Stefanie Weidtkamp-Peters, Marten Postma, Wenkun Zhou, Joachim Goedhart, María-Isabel Sánchez-Pérez, Theodorus W J Gadella, Rüdiger Simon, Ben Scheres, Ikram Blilou
During multicellular development, specification of distinct cell fates is often regulated by the same transcription factors operating differently in distinct cis-regulatory modules, either through different protein complexes, conformational modification of protein complexes, or combinations of both. Direct visualization of different transcription factor complex states guiding specific gene expression programs has been challenging. Here we use in vivo FRET-FLIM (Förster resonance energy transfer measured by fluorescence lifetime microscopy) to reveal spatial partitioning of protein interactions in relation to specification of cell fate...
July 26, 2017: Nature
Peng Zheng, Nianqiang Wu
Graphene oxide and graphene quantum dots are attractive fluorophores, which are inexpensive, non-toxic, photo-stable, water-soluble, biocompatible and environmentally friendly. They find extensive applications in fluorescent biosensors and chemi-sensors, in which they either serve as fluorophores or quenchers. As fluorophores, they display the tunable photoluminescence emission and the "Giant Red-Edge Effect". As quenchers, they exhibit a remarkable quenching efficiency via either electron transfer or Förster resonance energy transfer (FRET) process...
July 25, 2017: Chemistry, An Asian Journal
Huan Wang, Jeanne E Pemberton
The effect of solvent quality on the slip flow penetration of polymer films was evaluated by monitoring small-molecule mass transport under varying laminar flow rates using Förster resonance energy transfer in combination with total internal reflectance fluorescence microscopy (FRET-TIRFM). For thin films of poly(N-isopropylacrylamide) (pNIPAM), solvents with solvent quality ranging from good to poor were studied. The solvents used were composed of varying mole ratios of methanol and water in order to take advantage of the unique cononsolvency phenomenon of pNIPAM such that differences in the physicochemical properties of these solvents were insignificant for fluorescence detection...
July 24, 2017: Langmuir: the ACS Journal of Surfaces and Colloids
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