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Florian Richter, Ines Fonfara, Boris Bouazza, Charlotte Helene Schumacher, Majda Bratovič, Emmanuelle Charpentier, Andreas Möglich
Sensory photoreceptors have enabled non-invasive and spatiotemporal control of numerous biological processes. Photoreceptor engineering has expanded the repertoire beyond natural receptors, but to date no generally applicable strategy exists towards constructing light-regulated protein actuators of arbitrary function. We hence explored whether the homodimeric Rhodobacter sphaeroides light-oxygen-voltage (LOV) domain (RsLOV) that dissociates upon blue-light exposure can confer light sensitivity onto effector proteins, via a mechanism of light-induced functional site release...
October 15, 2016: Nucleic Acids Research
Sabyasachi Mukhopadhyay, Wolfgang Gärtner, David Cahen, Israel Pecht, Mordechai Sheves
Electron transport properties via a photochromic biological photoreceptor have been studied in junctions of monolayer assemblies in solid-state configurations. The photoreceptor studied was a member of the LOV domain protein family with a bound flavin chromophore, and its photochemically inactive mutant due to change of a crucial cysteine residue by a serine. The photochemical properties of the protein were maintained in dry, solid state conditions, indicating that the proteins in the junctions were assembled in native state-like conditions...
September 14, 2016: Physical Chemistry Chemical Physics: PCCP
Dimitris Petroutsos, Ryutaro Tokutsu, Shinichiro Maruyama, Serena Flori, Andre Greiner, Leonardo Magneschi, Loic Cusant, Tilman Kottke, Maria Mittag, Peter Hegemann, Giovanni Finazzi, Jun Minagawa
In plants and algae, light serves both as the energy source for photosynthesis and a biological signal that triggers cellular responses via specific sensory photoreceptors. Red light is perceived by bilin-containing phytochromes and blue light by the flavin-containing cryptochromes and/or phototropins (PHOTs), the latter containing two photosensory light, oxygen, or voltage (LOV) domains. Photoperception spans several orders of light intensity, ranging from far below the threshold for photosynthesis to values beyond the capacity of photosynthetic CO2 assimilation...
September 14, 2016: Nature
Aya Shimomura, Ayumi Naka, Nobuyuki Miyazaki, Sayaka Moriuchi, Susumu Arima, Shusei Sato, Hideki Hirakawa, Makoto Hayashi, Maskit Maymon, Ann M Hirsch, Akihiro Suzuki
In many legumes, roots that are exposed to light do not form nodules. Here, we report that blue light inhibits nodulation in Lotus japonicus roots inoculated with Mesorhizobium loti. Using RNA interference, we suppressed the expression of phototropin and cryptochrome genes in L. japonicus hairy roots. Under blue light, plants transformed with an empty vector did not develop nodules, whereas plants exhibiting suppressed expression of cry1 and cry2 genes formed nodules. We also measured rhizobial growth to investigate whether the inhibition of nodulation could be caused by a reduced population of rhizobia in response to light...
September 9, 2016: Molecular Plant-microbe Interactions: MPMI
Seth P Zimmerman, Ryan A Hallett, Ashley M Bourke, James E Bear, Matthew J Kennedy, Brian Kuhlman
Inducible dimers are powerful tools for controlling biological processes through colocalizing signaling molecules. To be effective, an inducible system should have a dissociation constant in the "off" state that is greater (i.e., weaker affinity) than the concentrations of the molecules that are being controlled, and in the "on" state a dissociation constant that is less (i.e., stronger affinity) than the relevant protein concentrations. Here, we reengineer the interaction between the light inducible dimer, iLID, and its binding partner SspB, to better control proteins present at high effective concentrations (5-100 μM)...
September 20, 2016: Biochemistry
Fanli Zhou, Karen M Qiang, Kathleen M Beckingham
The Drosophila protein Jim Lovell (Lov) is a putative transcription factor of the BTB/POZ (Bric- a-Brac/Tramtrack/Broad/ Pox virus and Zinc finger) domain class that is expressed in many elements of the developing larval nervous system. It has roles in innate behaviors such as larval locomotion and adult courtship. In performing tissue-specific knockdown with the Gal4-UAS system we identified a new behavioral phenotype for lov: larvae failed to burrow into their food during their growth phase and then failed to tunnel into an agarose substratum during their wandering phase...
2016: PloS One
Hui Wang, Marco Vilela, Andreas Winkler, Miroslaw Tarnawski, Ilme Schlichting, Hayretin Yumerefendi, Brian Kuhlman, Rihe Liu, Gaudenz Danuser, Klaus M Hahn
LOVTRAP is an optogenetic approach for reversible light-induced protein dissociation using protein A fragments that bind to the LOV domain only in the dark, with tunable kinetics and a >150-fold change in the dissociation constant (Kd). By reversibly sequestering proteins at mitochondria, we precisely modulated the proteins' access to the cell edge, demonstrating a naturally occurring 3-mHz cell-edge oscillation driven by interactions of Vav2, Rac1, and PI3K proteins.
September 2016: Nature Methods
Yuki Akiyama, Yusuke Nakasone, Yoichi Nakatani, Osamu Hisatomi, Masahide Terazima
Aureochrome (Aureo) is a recently discovered blue light sensor protein initially from Vaucheria frigida, in which it controls blue light-dependent branch formation and/or development of a sex organ by a light-dependent change in the affinity for DNA. Although photochemical reactions of Aureo-LOV (LOV is a C-terminal light-oxygen-voltage domain) and the N-terminal truncated construct containing a bZIP (N-terminal basic leucine zipper domain) and a LOV domain have previously been reported, the reaction kinetics of the change in affinity for DNA have never been elucidated...
August 4, 2016: Journal of Physical Chemistry. B
Katrin Röllen, Joachim Granzin, Vineet Panwalkar, Vladimir Arinkin, Raj Rani, Rudolf Hartmann, Ulrich Krauss, Karl-Erich Jaeger, Dieter Willbold, Renu Batra-Safferling
Light-Oxygen-Voltage (LOV) domains represent the photo-responsive domains of various blue-light photoreceptor proteins and are widely distributed in plants, algae, fungi, and bacteria. Here, we report the dark-state crystal structure of PpSB1-LOV, a slow-reverting short LOV protein from Pseudomonas putida that is remarkably different from our previously published "fully light-adapted" structure [1]. A direct comparison of the two structures provides insight into the light-activated signaling mechanism. Major structural differences involve a~11Å movement of the C terminus in helix Jα, ~4Å movement of Hβ-Iβ loop, disruption of hydrogen bonds in the dimer interface, and a~29° rotation of chain-B relative to chain-A as compared to the light-state dimer...
September 25, 2016: Journal of Molecular Biology
Jameela Lokhandwala, Rafael I Silverman Y de la Vega, Hilary C Hopkins, Collin W Britton, Aroa Rodriguez-Iglesias, Roberto Bogomolni, Monika Schmoll, Brian D Zoltowski
Light-oxygen-voltage (LOV) domain-containing proteins function as small light-activated modules capable of imparting blue light control of biological processes. Their small modular nature has made them model proteins for allosteric signal transduction and optogenetic devices. Despite intense research, key aspects of their signal transduction mechanisms and photochemistry remain poorly understood. In particular, ordered water has been identified as a possible key mediator of photocycle kinetics, despite the lack of ordered water in the LOV active site...
July 8, 2016: Journal of Biological Chemistry
Seokwoo Choi, Yusuke Nakasone, Klaas J Hellingwerf, Masahide Terazima
YtvA is a blue light sensor protein composed of an N-terminal LOV (light-oxygen-voltage) domain, a linker helix, and the C-terminal sulfate transporter and anti-σ factor antagonist domain. YtvA is believed to act as a positive regulator for light and salt stress responses by regulating the σB transcription factor. Although its biological function has been studied, the reaction dynamics and molecular mechanism underlying the function are not well understood. To improve our understanding of the signaling mechanism, we studied the reaction of the LOV domain (YLOV, amino acids 26-127), the LOV domain with its N-terminal extension (N-YLOV, amino acids 1-127), the LOV domain with its C-terminal linker helix (YLOV-linker, amino acids 26-147), and the YLOV domain with the N-terminal extension and the C-terminal linker helix (N-YLOV-linker, amino acids 1-147) using the transient grating method...
June 7, 2016: Biochemistry
Mengran Guo, Longshan Zhao, Mo Li, Qiang Fu, Xiaohui Pu, Bingyang Liu, Zhonggui He, Li Yang
Lovastatin (LOV) is an antihyperlipidemic agent which exhibits low bioavailability due to its poor solubility. Therefore, a nanosuspension (NS) was developed as an efficient strategy to improve its oral bioavailability. To evaluate the pharmacokinetics of LOV-NS, a novel, sensitive, and rapid UPLC-MS/MS method was developed and validated for the simultaneous determination of LOV and its metabolite lovastatin acid (LOVA) in rat plasma. Simvastatin (IS) was chosen as the internal standard, and a liquid-liquid extraction method was used to isolate LOV and LOVA from biological matrices...
June 15, 2016: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
Ankan Banerjee, Elena Herman, Manuel Serif, Manuel Maestre-Reyna, Sebastian Hepp, Richard Pokorny, Peter G Kroth, Lars-Oliver Essen, Tilman Kottke
The modular architecture of aureochrome blue light receptors, found in several algal groups including diatoms, is unique by having the LOV-type photoreceptor domain fused to the C-terminus of its putative effector, an N-terminal DNA-binding bZIP module. The structural and functional understanding of aureochromes' light-dependent signaling mechanism is limited, despite their promise as an optogenetic tool. We show that class I aureochromes 1a and 1c from the diatom Phaeodactylum tricornutum are regulated in a light-independent circadian rhythm...
July 8, 2016: Nucleic Acids Research
Michał Gabruk, Iwona Habina, Jerzy Kruk, Jolanta Dłużewska, Renata Szymańska
In this study, 25 accessions of Arabidopsis thaliana originating from a variety of climate conditions were grown under controlled circumstances of different light intensity and temperature. The accessions were analyzed for prenyllipids content and composition, as well as expression of the genes involved in tocochromanol biosynthesis (vte1-5). It was found that the applied conditions did not strongly affect total tocochromanols content and there was no apparent correlation of the tocochromanol content with the origin of the accessions...
June 2016: Physiologia Plantarum
Maxime M Grand, Petr Chocholouš, Jarda Růžička, Petr Solich, Christopher I Measures
By virtue of their compactness, long-term stability, minimal reagent consumption and robustness, miniaturized sequential injection instruments are well suited for automation of assays onboard research ships. However, in order to reach the sensitivity and limit of detection required for open-ocean determinations of trace elements, it is necessary to preconcentrate the analyte prior its derivatization and subsequent detection by fluorescence. In this work, a novel method for the determination of dissolved zinc (Zn) at subnanomolar levels in seawater is described...
June 7, 2016: Analytica Chimica Acta
Emanuel K Peter, Joan-Emma Shea, Igor V Pivkin
In this paper, we present a coarse replica exchange molecular dynamics (REMD) approach, based on kinetic Monte Carlo (kMC). The new development significantly can reduce the amount of replicas and the computational cost needed to enhance sampling in protein simulations. We introduce 2 different methods which primarily differ in the exchange scheme between the parallel ensembles. We apply this approach on folding of 2 different β-stranded peptides: the C-terminal β-hairpin fragment of GB1 and TrpZip4. Additionally, we use the new simulation technique to study the folding of TrpCage, a small fast folding α-helical peptide...
May 14, 2016: Physical Chemistry Chemical Physics: PCCP
Brian M Fuller, Ian Ferguson, Nicholas M Mohr, Robert J Stephens, Cristopher C Briscoe, Angelina A Kolomiets, Richard S Hotchkiss, Marin H Kollef
INTRODUCTION: In critically ill patients, acute respiratory distress syndrome (ARDS) and ventilator-associated conditions (VACs) are associated with increased mortality, survivor morbidity and healthcare resource utilisation. Studies conclusively demonstrate that initial ventilator settings in patients with ARDS, and at risk for it, impact outcome. No studies have been conducted in the emergency department (ED) to determine if lung-protective ventilation in patients at risk for ARDS can reduce its incidence...
2016: BMJ Open
Luana Marotta Reis de Vasconcellos, Mary Anne Moreira Barbara, Emanuel da Silva Rovai, Mariana de Oliveira França, Zahra Fernandes Ebrahim, Luis Gustavo Oliveira de Vasconcellos, Camila Deco Porto, Carlos Alberto Alves Cairo
The present study aimed to assess the effects of low-level laser therapy (GaAlAs) on the bone repair process within titanium scaffolds in the femurs of healthy and osteoporotic rats. Fifty-six rats were divided into four groups: group Sh: SHAM animals that received scaffolds; group LSh: SHAM animals that received scaffolds and were subjected to laser therapy; group OV: ovarietomized (OVX) animals that received scaffolds; and group LOV: OVX animals that received scaffolds and were subjected to laser therapy...
July 2016: Lasers in Medical Science
Rokhsana Yasmin, Shasha Rao, Kristen E Bremmell, Clive A Prestidge
Low dissolution of drugs in the intestinal fluid can limit their effectiveness in oral therapies. Here, a novel porous silica-supported solid lipid system was developed to optimize the oral delivery of drugs with limited aqueous solubility. Using lovastatin (LOV) as the model poorly water-soluble drug, two porous silica-supported solid lipid systems (SSL-A and SSL-S) were fabricated from solid lipid (glyceryl monostearate, GMS) and nanoporous silica particles Aerosil 380 (silica-A) and Syloid 244FP (silica-S) via immersion/solvent evaporation...
July 2016: AAPS Journal
Anthony M Buckley, Caitlin Jukes, Denise Candlish, June J Irvine, Janice Spencer, Robert P Fagan, Andrew J Roe, John M Christie, Neil F Fairweather, Gillian R Douce
The uses of fluorescent reporters derived from green fluorescent protein have proved invaluable for the visualisation of biological processes in bacteria grown under aerobic conditions. However, their requirement for oxygen has limited their application in obligate anaerobes such as Clostridium difficile. Fluorescent proteins derived from Light, Oxygen or Voltage sensing (LOV) domains have been shown to bridge this limitation, but their utility as translational fusions to monitor protein expression and localisation in a strict anaerobic bacterium has not been reported...
2016: Scientific Reports
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