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https://www.readbyqxmd.com/read/28293891/development-of-a-synthetic-switch-to-control-protein-stability-in-eukaryotic-cells-with-light
#1
Christof Taxis
In eukaryotic cells, virtually all regulatory processes are influenced by proteolysis. Thus, synthetic control of protein stability is a powerful approach to influence cellular behavior. To achieve this, selected target proteins are modified with a conditional degradation sequence (degron) that responds to a distinct signal. For development of a synthetic degron, an appropriate sensor domain is fused with a degron such that activity of the degron is under control of the sensor. This chapter describes the development of a light-activated, synthetic degron in the model organism Saccharomyces cerevisiae...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28271102/switching-from-adduct-formation-to-electron-transfer-in-a-light-oxygen-voltage-domain-containing-the-reactive-cysteine
#2
Kathrin Magerl, Ivan Stambolic, Bernhard Dick
LOV (light-, oxygen- or voltage-sensitive) domains act as photosensory units of many prokaryotic and eukaryotic proteins. Upon blue light excitation they undergo a photocycle via the excited triplet state of their flavin chromophore yielding the flavin-cysteinyl adduct. Adduct formation is highly conserved among all LOV domains and constitutes the primary step of LOV domain signaling. But recently, it has been shown that signal propagation can also be triggered by flavin photoreduction to the neutral semiquinone offering new prospects for protein engineering...
March 8, 2017: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/28259481/lung-protective-ventilation-initiated-in-the-emergency-department-lov-ed-a%C3%A2-quasi-experimental-before-after-trial
#3
Brian M Fuller, Ian T Ferguson, Nicholas M Mohr, Anne M Drewry, Christopher Palmer, Brian T Wessman, Enyo Ablordeppey, Jacob Keeperman, Robert J Stephens, Cristopher C Briscoe, Angelina A Kolomiets, Richard S Hotchkiss, Marin H Kollef
STUDY OBJECTIVE: We evaluated the efficacy of an emergency department (ED)-based lung-protective mechanical ventilation protocol for the prevention of pulmonary complications. METHODS: This was a quasi-experimental, before-after study that consisted of a preintervention period, a run-in period of approximately 6 months, and a prospective intervention period. The intervention was a multifaceted ED-based mechanical ventilator protocol targeting lung-protective tidal volume, appropriate setting of positive end-expiratory pressure, rapid oxygen weaning, and head-of-bed elevation...
March 1, 2017: Annals of Emergency Medicine
https://www.readbyqxmd.com/read/28257211/temperature-sensitive-singlet-oxygen-photosensitization-by-lov-derived-fluorescent-flavoproteins
#4
Michael Westberg, Mikkel Bregnhøj, Michael Etzerodt, Peter R Ogilby
Optogenetic sensitizers that selectively produce a given reactive oxygen species (ROS) constitute a promising tool for studying cell signaling processes with high levels of spatiotemporal control. However, to harness the full potential of this tool for live cell studies, the photophysics of currently available systems need to be explored further and optimized. Of particular interest in this regard, are the flavoproteins miniSOG and SOPP, both of which (1) contain the chromophore flavin mononucleotide, FMN, in a LOV-derived protein enclosure, and (2) photosensitize the production of singlet oxygen, O2(a(1)Δg)...
March 16, 2017: Journal of Physical Chemistry. B
https://www.readbyqxmd.com/read/28244872/kinetics-of-the-lov-domain-of-zeitlupe-determine-its-circadian-function-in-arabidopsis
#5
Ashutosh Pudasaini, Jae Sung Shim, Young Hun Song, Hua Shi, Takatoshi Kiba, David E Somers, Takato Imaizumi, Brian D Zoltowski
A LOV (Light, Oxygen, or Voltage) domain containing blue-light photoreceptor ZEITLUPE (ZTL) directs circadian timing by degrading clock proteins in plants. Functions hinge upon allosteric differences coupled to the ZTL photocycle; however, structural and kinetic information was unavailable. Herein, we tune the ZTL photocycle over two orders of magnitude. These variants reveal that ZTL complexes with targets independent of light, but dictates enhanced protein degradation in the dark. In vivo experiments definitively show photocycle kinetics dictate the rate of clock component degradation, thereby impacting circadian period...
February 28, 2017: ELife
https://www.readbyqxmd.com/read/28229215/sequence-analysis-and-expression-of-a-blue-light-photoreceptor-gene-slwc-1-from-the-cauliflower-mushroom-sparassis-latifolia
#6
Chi Yang, Lu Ma, Zhenghe Ying, Xiaoling Jiang, Yanquan Lin
Light is a necessary environmental factor for fruit body formation and development of the cauliflower mushroom Sparassis latifolia, a well-known edible and medicinal fungus. In this study, we firstly characterized the SP-C strain, which belonged to S. latifolia. And then we cloned and sequenced a photoreceptor gene (Slwc-1) from S. latifolia. The product of Slwc-1, SlWC-1 (872 aa residues) contained a coiled-coil region, a LOV domain, and two PAS domains. Phylogenetic tree result showed that SLWC-1 was most close to GfWC-1 from Grifola frondosa in edible and medicinal fungus...
April 2017: Current Microbiology
https://www.readbyqxmd.com/read/28211532/structure-of-a-lov-protein-in-apo-state-and-implications-for-construction-of-lov-based-optical-tools
#7
Vladimir Arinkin, Joachim Granzin, Katrin Röllen, Ulrich Krauss, Karl-Erich Jaeger, Dieter Willbold, Renu Batra-Safferling
Unique features of Light-Oxygen-Voltage (LOV) proteins like relatively small size (~12-19 kDa), inherent modularity, highly-tunable photocycle and oxygen-independent fluorescence have lately been exploited for the generation of optical tools. Structures of LOV domains reported so far contain a flavin chromophore per protein molecule. Here we report two new findings on the short LOV protein W619_1-LOV from Pseudomonas putida. First, the apo-state crystal structure of W619_1-LOV at 2.5 Å resolution reveals conformational rearrangements in the secondary structure elements lining the chromophore pocket including elongation of the Fα helix, shortening of the Eα-Fα loop and partial unfolding of the Eα helix...
February 17, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28194625/trypanosomiasis-challenge-estimation-using-the-diminazene-aceturate-berenil-index-in-zebu-in-gabon
#8
Brieuc G A Cossic, Brice Adjahoutonon, Pierre Gloaguen, Gui Lov Dibanganga, Gael Maganga, Pascal Leroy, Ewan T MacLeod, Kim Picozzi
A longitudinal study was conducted within a cattle ranch in Gabon to determine the diminazene aceturate (Berenil) index (DAI) in a group of Zebu, raised under low tsetse density; this measure providing an assessment of trypanosomiasis risk. The objective was to evaluate the trypanosomiasis pressure thus informing trypanosomiasis control methods and cattle management. Twenty female adult Zebu were monitored for 24 weeks during the dry season. Blood samples were collected on a weekly basis and subjected to parasitological and haematological analysis (n = 480), using the buffy-coat method and the packed cell volume value (PCV), respectively, infected animals were treated with a single intramuscular injection of diminazene aceturate (8 mg/kg)...
March 2017: Tropical Animal Health and Production
https://www.readbyqxmd.com/read/28145707/glutamine-amide-flip-elicits-long-distance-allosteric-responses-in-the-lov-protein-vivid
#9
Abir Ganguly, Walter Thiel, Brian R Crane
Light-oxygen-voltage (LOV) domains sense blue light through the photochemical formation of a cysteinyl-flavin covalent adduct. Concurrent protonation at the flavin N5 position alters the hydrogen bonding interactions of an invariant Gln residue that has been proposed to flip its amide side chain as a critical step in the propagation of conformational change. Traditional molecular dynamics (MD) and replica-exchange MD (REMD) simulations of the well-characterized LOV protein Vivid (VVD) demonstrate that the Gln182 amide indeed reorients by ∼180° in response to either adduct formation or reduction of the isoalloxazine ring to the neutral semiquinone, both of which involve N5 protonation...
February 13, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28127721/stabilization-and-amorphization-of-lovastatin-using-different-types-of-silica
#10
Mai Khanfar, Suhair Al-Nimry
Lovastatin (LOV), an antihyperlipidimic agent, is characterized by low solubility/poor dissolution and, thus, low bioavailability (<5%). A beneficial effect on its bioavailability could result from improving its dissolution. One of the most common methods used to enhance dissolution is the preparation of solid dispersions. Solid dispersions of LOV and silica with different surface areas were prepared. The effects of the type of silica, ratio of drug/silica, incubation period with silica, and the effect of surface area were all studied...
January 26, 2017: AAPS PharmSciTech
https://www.readbyqxmd.com/read/28100053/the-grateful-infrared-sequential-protein-structural-changes-resolved-by-infrared-difference-spectroscopy
#11
Tilman Kottke, Víctor A Lórenz-Fonfría, Joachim Heberle
The catalytic activity of proteins is a function of structural changes. Very often these are as minute as protonation changes, hydrogen bonding changes, and amino acid side chain reorientations. To resolve these, a methodology is afforded that not only provides the molecular sensitivity but allows for tracing the sequence of these hierarchical reactions at the same time. This feature article showcases results from time-resolved IR spectroscopy on channelrhodopsin (ChR), light-oxygen-voltage (LOV) domain protein, and cryptochrome (CRY)...
January 19, 2017: Journal of Physical Chemistry. B
https://www.readbyqxmd.com/read/28068090/femtosecond-to-millisecond-dynamics-of-light-induced-allostery-in-the-avena-sativa-lov-domain
#12
Agnieszka A Gil, Sergey P Laptenok, Jarrod B French, James N Iuliano, Andras Lukacs, Christopher R Hall, Igor V Sazanovich, Gregory M Greetham, Adelbert Bacher, Boris Illarionov, Markus Fischer, Peter J Tonge, Stephen R Meech
The rational engineering of photosensor proteins underpins the field of optogenetics, in which light is used for spatiotemporal control of cell signaling. Optogenetic elements function by converting electronic excitation of an embedded chromophore into structural changes on the microseconds to seconds time scale, which then modulate the activity of output domains responsible for biological signaling. Using time-resolved vibrational spectroscopy coupled with isotope labeling, we have mapped the structural evolution of the LOV2 domain of the flavin binding phototropin Avena sativa (AsLOV2) over 10 decades of time, reporting structural dynamics between 100 fs and 1 ms after optical excitation...
February 9, 2017: Journal of Physical Chemistry. B
https://www.readbyqxmd.com/read/28055118/fret-in-a-synthetic-flavin-and-bilin-binding-protein
#13
Julian Simon, Aba Losi, Kai-Hong Zhao, Wolfgang Gärtner
The last decade has seen development and application of a large number of novel fluorescence-based techniques that have revolutionized fluorescence microscopy in life sciences. Preferred tags for such applications are genetically encoded fluorescent proteins (FP), mostly derivatives of the green fluorescent protein (GFP). Combinations of FPs with wavelength-separated absorption/fluorescence properties serve as excellent tools for molecular interaction studies, e.g., protein-protein complexes or enzyme-substrate complexes, based on the FRET phenomenon (Förster resonance energy transfer)...
January 5, 2017: Photochemistry and Photobiology
https://www.readbyqxmd.com/read/27906450/lovtrap-a-versatile-method-to-control-protein-function-with-light
#14
Hui Wang, Klaus M Hahn
We describe a detailed procedure for the use of LOVTRAP, an approach to reversibly sequester and release proteins from cellular membranes using light. In the application described here, proteins that act at the plasma membrane are held at mitochondria in the dark, and reversibly released by irradiation. The technique relies on binding of an engineered Zdk domain to a LOV2 domain, with affinity <30 nM in the dark and >500 nM upon irradiation between 400 and 500 nm. LOVTRAP can be applied to diverse proteins, as it requires attaching only one member of the Zdk/LOV2 pair to the target protein, and the other to the membrane where the target protein is to be sequestered...
December 1, 2016: Current Protocols in Cell Biology
https://www.readbyqxmd.com/read/27861974/solving-blue-light-riddles-new-lessons-from-flavin-binding-lov-photoreceptors
#15
REVIEW
Aba Losi, Wolfgang Gärtner
Detection of blue light (BL) via flavin-binding photoreceptors (Fl-Blues) has evolved throughout all three domains of life. Although the main BL players, that is light, oxygen and voltage (LOV), blue light sensing using flavins (BLUF) and Cry (cryptochrome) proteins, have been characterized in great detail with respect to structure and function, still several unresolved issues at different levels of complexity remain and novel unexpected findings were reported. Here, we review the most prevailing riddles of LOV-based photoreceptors, for example: the relevance of water and/or small metabolites for the dynamics of the photocycle; molecular details of light-to-signal transduction events; the interplay of BL sensing by LOV domains with other environmental stimuli, such as BL plus oxygen-mediating photodamage and its impact on microbial lifestyles; the importance of the cell or chromophore redox state in determining the fate of BL-driven reactions; the evolutionary pathways of LOV-based BL sensing and associated functions through the diverse phyla...
November 18, 2016: Photochemistry and Photobiology
https://www.readbyqxmd.com/read/27849731/51-emergency-department-lung-protective-ventilation-reduces-pulmonary-complications-the-lov-ed-trial
#16
Brian Fuller, Ian Ferguson, Christopher Palmer, Anne Drewry, Brian Wessman, Enyo Ablordeppey, Nicholas Mohr, Marin Kollef
No abstract text is available yet for this article.
December 2016: Critical Care Medicine
https://www.readbyqxmd.com/read/27832823/resting-energy-expenditure-calorie-and-protein-consumption-in-critically-ill-patients-a-retrospective-cohort-study
#17
Oren Zusman, Miriam Theilla, Jonathan Cohen, Ilya Kagan, Itai Bendavid, Pierre Singer
BACKGROUND: Intense debate exists regarding the optimal energy and protein intake for intensive care unit (ICU) patients. However, most studies use predictive equations, demonstrated to be inaccurate to target energy intake. We sought to examine the outcome of a large cohort of ICU patients in relation to the percent of administered calories divided by resting energy expenditure (% AdCal/REE) obtained by indirect calorimetry (IC) and to protein intake. METHODS: Included patients were hospitalized from 2003 to 2015 at a 16-bed ICU at a university affiliated, tertiary care hospital, and had IC measurement to assess caloric targets...
November 10, 2016: Critical Care: the Official Journal of the Critical Care Forum
https://www.readbyqxmd.com/read/27744350/engineering-of-temperature-and-light-switchable-cas9-variants
#18
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...
November 16, 2016: Nucleic Acids Research
https://www.readbyqxmd.com/read/27722652/electron-transport-via-a-soluble-photochromic-photoreceptor
#19
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
https://www.readbyqxmd.com/read/27626383/a-blue-light-photoreceptor-mediates-the-feedback-regulation-of-photosynthesis
#20
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 22, 2016: Nature
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