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Dieter Oesterhelt - One of the best experts on this subject based on the ideXlab platform.
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a small basic protein from the brz brb operon is involved in regulation of bop transcription in halobacterium salinarum
BMC Molecular Biology, 2011Co-Authors: Valery Tarasov, Rita Schwaiger, Katarina Furtwangler, Mike Dyallsmith, Dieter OesterheltAbstract:Background The halophilic archaeon Halobacterium salinarum expresses Bacteriorhodopsin, a retinal-protein that allows photosynthetic growth. Transcription of the bop (b acterioop sin) gene is controlled by two transcription factors, Bat and Brz that induce bop when cells are grown anaerobically and under light.
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primary reaction of sensory rhodopsin ii mutant d75n and the influence of azide
Biochemistry, 2009Co-Authors: Mirka-kristin Verhoefen, Johann P. Klare, Martin Engelhard, Sergiu Amarie, Martin O. Lenz, Jörg Tittor, Dieter Oesterhelt, Josef WachtveitlAbstract:The early steps in the photocycle of sensory rhodopsin II mutant D75N are investigated in a comprehensive study using femtosecond visible pump/probe spectroscopy. An overall slower response dynamics after photoexcitation is observed compared to wild-type sensory rhodopsin II, which is assigned to changed electrostatics and an altered hydrogen-bonding network within the retinal binding pocket. Furthermore, the influence of azide on the primary reaction is analyzed. The addition of azide accelerates the sub-10 ps dynamics of the D75N mutant nearly to reaction rates found in wild-type. Moreover, a further reaction pathway becomes observable in the investigated time range, which is assigned to a previously described K1 to K2 transition. The specific acceleration of the early steps seems to be a unique feature of the D75N mutant as similar azide effects do not emerge in analogous azide measurements of wild-type sensory rhodopsin II, Bacteriorhodopsin, and the Bacteriorhodopsin mutant D85N.
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a small protein from the bop brp intergenic region of halobacterium salinarum contains a zinc finger motif and regulates bop and crtb1 transcription
Molecular Microbiology, 2008Co-Authors: Valery Tarasov, Rita Schwaiger, Katarina Furtwangler, Huseyin Besir, Kathrin Klee, Friedhelm Pfeiffer, Dieter OesterheltAbstract:Bacteriorhodopsin, the photosynthetic protein of Halobacterium salinarum, is optimally expressed under anaerobic growth conditions. We identified Brz (OE3104F, Bacteriorhodopsin-regulating zinc finger protein), a new regulator of the bop gene. It is a small protein with a zinc finger motif, encoded directly upstream of the bop gene in the same orientation. Deletion of the brz gene caused a large decrease of bop mRNA levels as shown by Northern blot and microarray analysis. A similar effect was obtained by site-directed mutagenesis of Cys and His residues in the zinc finger motif, indicating the importance of this motif for the function of the protein. In silico analysis of the genomes from H. salinarum and other archaea revealed a large family of similar small zinc finger motif proteins, some of which may also be involved in transcription regulation of their adjacent genes.
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in situ determination of transient pka changes of internal amino acids of Bacteriorhodopsin by using time resolved attenuated total reflection fourier transform infrared spectroscopy
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Christian Zscherp, Jörg Tittor, Dieter Oesterhelt, Ramona Schlesinger, Joachim HeberleAbstract:Active proton transfer through membrane proteins is accomplished by shifts in the acidity of internal amino acids, prosthetic groups, and water molecules. The recently introduced step-scan attenuated total reflection Fourier-transform infrared (ATR/FT-IR) spectroscopy was employed to determine transient pKa changes of single amino acid side chains of the proton pump Bacteriorhodopsin. The high pKa of D96 (>12 in the ground state) drops to 7.1 +/- 0.2 (in 1 M KCl) during the lifetime of the N intermediate, quantitating the role of D96 as the internal proton donor of the retinal Schiff base. We conclude from experiments on the pH dependence of the proton release reaction and on point mutants where each of the glutamates on the extracellular surface has been exchanged that besides D85 no other carboxylic group changes its protonation state during proton release. However, E194 and E204 interact with D85, the primary proton acceptor of the Schiff base proton. The C==O stretching vibration of D85 undergoes a characteristic pH-dependent shift in frequency during the M state of wild-type Bacteriorhodopsin with a pKa of 5.2 (+/-0.3) which is abolished in the single-site mutants E194Q and E204Q and the quadruple mutant E9Q/E74Q/E194Q/E204Q. The double mutation E9Q/E74Q does not affect the lifetime of the intermediates, ruling out any participation of these residues in the proton transfer chain of Bacteriorhodopsin. This study demonstrates that transient changes in acidity of single amino acid residues can be quantified in situ with infrared spectroscopy.
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localization of glycolipids in membranes by in vivo labeling and neutron diffraction
Molecular Cell, 1998Co-Authors: Martin Weik, Heiko Patzelt, Giuseppe Zaccai, Dieter OesterheltAbstract:Abstract Evidence is accumulating for the lateral organization of cell membrane lipids and proteins in the context of sorting or intracellular signaling. So far, however, information has been lacking on the details of protein–lipid interactions in such aggregates. Purple membranes are patches made up of lipids and the protein Bacteriorhodopsin in the plasma membrane of certain Archaea. Naturally crystalline, they provide a unique opportunity to study the structure of a natural membrane at submolecular resolution by diffraction methods. We present a direct structural determination of the glycolipids with respect to Bacteriorhodopsin in these membranes. Deuterium labels incorporated in vivo into the sugar moieties of the major glycolipid were localized by neutron diffraction. The data suggest a role for specific aromatic residue–carbohydrate stacking interactions in the formation of the purple membrane crystalline patches.
Roland R Netz - One of the best experts on this subject based on the ideXlab platform.
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Orientation of non-spherical protonated water clusters revealed by infrared absorption dichroism
Nature Communications, 2018Co-Authors: Jan O. Daldrop, Mattia Saita, Víctor A. Lórenz-fonfría, Matthias Heyden, Johannes Heberle, Roland R NetzAbstract:Infrared continuum bands that extend over a broad frequency range are a key spectral signature of protonated water clusters. They are observed for many membrane proteins that contain internal water molecules, but their microscopic mechanism has remained unclear. Here we compute infrared spectra for protonated and unprotonated water chains, discs, and droplets from ab initio molecular dynamics simulations. The continuum bands of the protonated clusters exhibit significant anisotropy for chains and discs, with increased absorption along the direction of maximal cluster extension. We show that the continuum band arises from the nuclei motion near the excess charge, with a long-ranged amplification due to the electronic polarizability. Our experimental, polarization-resolved light–dark difference spectrum of the light-driven proton pump Bacteriorhodopsin exhibits a pronounced dichroic continuum band. Our results suggest that the protonated water cluster responsible for the continuum band of Bacteriorhodopsin is oriented perpendicularly to the membrane normal.Protein-bound water clusters play a key role for proton transport and storage in molecular biology. Here, the authors show by simulations and experiments that the orientation of non-spherical protonated water clusters in Bacteriorhodopsin is unveiled by polarization-resolved infrared spectroscopy.
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Orientation of non-spherical protonated water clusters revealed by infrared absorption dichroism
Nature Publishing Group, 2018Co-Authors: Jan O. Daldrop, Mattia Saita, Víctor A. Lórenz-fonfría, Matthias Heyden, Joachim Heberle, Roland R NetzAbstract:Protein-bound water clusters play a key role for proton transport and storage in molecular biology. Here, the authors show by simulations and experiments that the orientation of non-spherical protonated water clusters in Bacteriorhodopsin is unveiled by polarization-resolved infrared spectroscopy
Gyorgy Varo - One of the best experts on this subject based on the ideXlab platform.
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characterization of the proton transporting photocycle of pharaonis halorhodopsin
Biophysical Journal, 2000Co-Authors: Agnes Kulcsar, Geza I Groma, Gyorgy VaroAbstract:The photocycle of pharaonis halorhodopsin was investigated in the presence of 100 mM NaN(3) and 1 M Na(2)SO(4). Recent observations established that the replacement of the chloride ion with azide transforms the photocycle from a chloride-transporting one into a proton-transporting one. Kinetic analysis proves that the photocycle is very similar to that of Bacteriorhodopsin. After K and L, intermediate M appears, which is missing from the chloride-transporting photocycle. In this intermediate the retinal Schiff base deprotonates. The rise of M in halorhodopsin is in the microsecond range, but occurs later than in Bacteriorhodopsin, and its decay is more accentuated multiphasic. Intermediate N cannot be detected, but a large amount of O accumulates. The multiphasic character of the last step of the photocycle could be explained by the existence of a HR' state, as in the chloride photocycle. Upon replacement of chloride ion with azide, the fast electric signal changes its sign from positive to negative, and becomes similar to that detected in Bacteriorhodopsin. The photocycle is enthalpy-driven, as is the chloride photocycle of halorhodopsin. These observations suggest that, while the basic charge translocation steps become identical to those in Bacteriorhodopsin, the storage and utilization of energy during the photocycle remains unchanged by exchanging chloride with azide.
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analogies between halorhodopsin and Bacteriorhodopsin
Biochimica et Biophysica Acta, 2000Co-Authors: Gyorgy VaroAbstract:The light-activated proton-pumping Bacteriorhodopsin and chloride ion-pumping halorhodopsin are compared. They belong to the family of retinal proteins, with 25% amino acid sequence homology. Both proteins have seven α helices across the membrane, surrounding the retinal binding pocket. Photoexcitation of all-trans retinal leads to ion transporting photocycles, which exhibit great similarities in the two proteins, despite the differences in the ion transported. The spectra of the K, L, N and O intermediates, calculated using time-resolved spectroscopic measurements, are very similar in both proteins. The absorption kinetic measurements reveal that the chloride ion transporting photocycle of halorhodopsin does not have intermediate M characteristic for deprotonated Schiff base, and intermediate L dominates the process. Energetically the photocycle of Bacteriorhodopsin is driven mostly by the decrease of the entropic energy, while the photocycle of halorhodopsin is enthalpy-driven. The ion transporting steps were characterized by the electrogenicity of the intermediates, calculated from the photoinduced transient electric signal measurements. The function of both proteins could be described with the ‘local access’ model developed for Bacteriorhodopsin. In the framework of this model it is easy to understand how Bacteriorhodopsin can be converted into a chloride pump, and halorhodopsin into a proton pump, by changing the ion specificity with added ions or site-directed mutagenesis.
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light driven chloride ion transport by halorhodopsin from natronobacterium pharaonis 1 the photochemical cycle
Biochemistry, 1995Co-Authors: Gyorgy Varo, Hideki Kandori, Leonid S. Brown, Akio Maeda, Jun Sasaki, Richard NeedlemanAbstract:The photochemical cycle of the light-driven chloride pump, halorhodopsin from N. pharonis, is described by transient optical multichannel and single-wavelength spectroscopy in the visible, and in the infrared. Titration of a blue-shift of the adsorption maximum upon addition of chloride describes a binding site with a K{sub D} of 1 mM. The reaction sequence after the all-trans to 13-cis photoisomerization of the retinal in this chloride binding form is itself dependent on the chloride. At 2 M chloride it is described by the scheme: HR {yields} K {leftrightarrow} L {leftrightarrow} N {yields} HR that relaxes in a few milliseconds, and is very similar to the photocycle of Bacteriorhodopsin under conditions where the retinal Schiff base cannot deprotonate. At lower chloride concentrations, e.g., 0.1 M, however, a red-shifted state termed O appears between N and HR, in equilibrium with N. The absorption spectra of K, L, N, and O are very similiar to their counterparts in the Bacteriorhodopsin photocycle. As in their equivalents in Bacteriorhodopsin, in the N state the retinal is still 13-cis, but it is reisomerized in the O state to all-trans. 90 refs., 8 figs., 1 tab.
Jan O. Daldrop - One of the best experts on this subject based on the ideXlab platform.
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Orientation of non-spherical protonated water clusters revealed by infrared absorption dichroism
Nature Communications, 2018Co-Authors: Jan O. Daldrop, Mattia Saita, Víctor A. Lórenz-fonfría, Matthias Heyden, Johannes Heberle, Roland R NetzAbstract:Infrared continuum bands that extend over a broad frequency range are a key spectral signature of protonated water clusters. They are observed for many membrane proteins that contain internal water molecules, but their microscopic mechanism has remained unclear. Here we compute infrared spectra for protonated and unprotonated water chains, discs, and droplets from ab initio molecular dynamics simulations. The continuum bands of the protonated clusters exhibit significant anisotropy for chains and discs, with increased absorption along the direction of maximal cluster extension. We show that the continuum band arises from the nuclei motion near the excess charge, with a long-ranged amplification due to the electronic polarizability. Our experimental, polarization-resolved light–dark difference spectrum of the light-driven proton pump Bacteriorhodopsin exhibits a pronounced dichroic continuum band. Our results suggest that the protonated water cluster responsible for the continuum band of Bacteriorhodopsin is oriented perpendicularly to the membrane normal.Protein-bound water clusters play a key role for proton transport and storage in molecular biology. Here, the authors show by simulations and experiments that the orientation of non-spherical protonated water clusters in Bacteriorhodopsin is unveiled by polarization-resolved infrared spectroscopy.
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Orientation of non-spherical protonated water clusters revealed by infrared absorption dichroism
Nature Publishing Group, 2018Co-Authors: Jan O. Daldrop, Mattia Saita, Víctor A. Lórenz-fonfría, Matthias Heyden, Joachim Heberle, Roland R NetzAbstract:Protein-bound water clusters play a key role for proton transport and storage in molecular biology. Here, the authors show by simulations and experiments that the orientation of non-spherical protonated water clusters in Bacteriorhodopsin is unveiled by polarization-resolved infrared spectroscopy
Robert M Glaeser - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of the bromide bound d85s mutant of Bacteriorhodopsin principles of ion pumping
Biophysical Journal, 2003Co-Authors: Marc T Facciotti, Vincent S Cheung, Doris Nguyen, Shahab Rouhani, Robert M GlaeserAbstract:We report the crystal structure of a bromide-bound form of the D85S mutant of Bacteriorhodopsin, bR(D85S), a protein that uses light energy rather than ATP to pump halide ions across the cell membrane. Comparison of the structure of the halide-bound and halide-free states reveals that both displacements of individual side-chain positions and concerted helical movements occur on the extracellular side of the protein. Analysis of these structural changes reveals how this ion pump first facilitates ion uptake deep within the cell membrane and then prevents the backward escape of ions later in the pumping cycle. Together with the information provided by structures of intermediate states in the Bacteriorhodopsin photocycle, this study also suggests the overall design principles that are necessary for ion pumping.
