Optical control of the primary step photoisomerization retinal molecule in bacteriorhodopsin from all-trans to 13-cis state was demonstrated under weak field conditions (where only 1 300 molecules absorbs a photon during excitation cycle) that are relevant understanding biological processes. By modulating phases and amplitudes spectral components photoexcitation pulse, we showed absolute quantity formed upon can be enhanced or suppressed by ±20% yield observed using short transform-limited...

We have measured proton release into the medium after transfer from retinal Schiff base to Asp85 in photocycle and C=O stretch bands of carboxylic acids wild type bacteriorhodopsin E204Q E204D mutants. In E204Q, but not E204D, normal is absent. Consistent with this, a negative band Fourier transform infrared difference spectra at 1700 cm-1 type, which we now attribute depletion protonated E204, also absent E204Q. this shifted 1714 cm-1, as expected higher frequency for aspartic than glutamic...

In the light-driven proton pump bacteriorhodopsin, transfer from retinal Schiff base to aspartate-85 is crucial reaction of transport cycle. halorhodopsin, a chloride ion pump, equivalent residue 85 threonine. When was replaced with threonine, mutated bacteriorhodopsin became when expressed in Halobacterium salinarium and, like actively transported ions direction opposite pump. Chloride bound it, as revealed by large shifts absorption maximum chromophore, and its photointermediates included...

Bacteriorhodopsin-like proteins provide archaea and eubacteria with a unique bioenergetic pathway comprising light-driven transmembrane proton translocation by single retinal-binding protein. Recently, homologous were found to perform photosensory functions in lower eukaryotes, but no active ion transport eukaryotic rhodopsins was detected. By demonstrating pumping fungal rhodopsin from Leptosphaeria maculans , we present case of retinal-based transporter eukaryote. This result implies that...

The spectral and photochemical properties of proteorhodopsin (PR) were determined to compare its proton transport steps those bacteriorhodopsin (BR). Static time-resolved measurements on wild-type PR several mutants done in the visible infrared (FTIR FT-Raman). Assignment observed CO stretch bands indicated that Asp-97 Glu-108 serve as acceptor donor, respectively, retinal Schiff base, do residues at corresponding positions BR, but there are numerous kinetic differences between two proteins....

Proton transfer is crucial for many enzyme reactions. Here, we show that in addition to protonatable amino acid side chains, water networks could constitute proton-binding sites proteins. A broad IR continuum absorbance change during the proton pumping photocycle of bacteriorhodopsin (bR) indicates most likely deprotonation a protonated cluster at release site close surface. We investigate influence several mutations on network and change, gain information about location extent reveal...

Glu-194 near the extracellular surface of bacteriorhodopsin is indispensable for proton release to medium upon protonation Asp-85 during light-driven transport. As Glu-204, its replacement with glutamine (but not aspartate) abolishes both and anomalous titration that originates from coupling between pKa this buried aspartate those other acidic groups. Unlike case however, raises release. In Fourier transform infrared spectra E194D mutant a prominent positive band observed at 1720 cm-1. It...

Because asp-85 is the acceptor of retinal Schiff base proton during light-driven transport by bacteriorhodopsin, modulation its pKa in photocycle to be expected. The complex titration unphotolyzed protein was suggested [Balashov, S. P., Govindjee, R., Imasheva, E. S., Misra, Ebrey, T. G., Feng, Y., Crouch, R. K., & Menick, D. R (1995) Biochemistry 34, 8820−8834] reflect dependence this residue on protonation state another, unidentified group. From pH dependencies rate constant for thermal...

Break on through: High-resolution multidimensional solid-state NMR spectroscopy was used to refine the molecular conformation of a seven-helical transmembrane photoreceptor in lipids (see picture, right). H/D exchange experiments left; black spectrum: H2O, red D2O) reveal solvent-exposed surface protein, which is asymmetrically positioned bilayer.

We used high-resolution proton-detected multidimensional NMR to study the solvent-exposed parts of a seven-helical integral membrane proton pump, proteorhodopsin (PR). PR samples were prepared by growing apoprotein on fully deuterated medium and reintroducing protons solvent-accessible sites through exchange with protonated buffer. This preparation leads spectra resolution down ca. 0.2 ppm at fast spinning (28 kHz) in protein back-exchanged level 40%. Novel three-dimensional chemical shift...

The ability to detect and characterize molecular motions represents one of the unique strengths nuclear magnetic resonance (NMR) spectroscopy. In this study, we report solid-state NMR site-specific measurements dipolar order parameters (15)N rotating frame spin-lattice (R1ρ) relaxation rates in a seven transmembrane helical protein Anabaena Sensory Rhodopsin reconstituted lipids. magnitudes observed indicate that both well-defined regions less structured intramembrane loops undergo...

Protein-protein interactions play critical roles in cellular function and oligomerization of membrane proteins is a commonly observed phenomenon. Determining the state defining intermolecular interface bilayer generally difficult task. Here, we use site-specific spin labeling to demonstrate that relaxation enhancements induced by covalently attached paramagnetic tag can provide distance restraints intermonomer oligomers formed seven-helical transmembrane protein Anabaena Sensory Rhodopsin...

Photoactivated adenylyl cyclase (PAC) and guanylyl rhodopsin increase the concentrations of intracellular cyclic nucleotides upon illumination, serving as promising second-generation tools in optogenetics. To broaden arsenal such tools, it is desirable to have light-activatable enzymes that can decrease nucleotide cells. Here, we report on an unusual microbial may be able meet demand. It found choanoflagellate Salpingoeca rosetta contains a C-terminal phosphodiesterase (PDE) domain. We...

Significance Our identification and characterization of two anion channelrhodopsins families, one from nonphotosynthetic microorganisms, shows that light-gated conductance is more widely spread among eukaryotic lineages than previously thought. The strongly red-shifted absorption spectra the subset we designate RubyACRs make them promising candidates for long-sought inhibitory optogenetic tools producing large passive currents activated by red light, enabling deep tissue penetration....

Abstract Gloeobacter rhodopsin (GR) is a cyanobacterial proton pump which can be potentially applied to optogenetics. We solved the crystal structure of GR and found that it has overall similarity homologous from Salinibacter ruber , xanthorhodopsin (XR). identified distinct structural characteristics GR’s hydrogen bonding network in transmembrane domain as well displacement extracellular sides helices relative those XR. Employing Raman spectroscopy flash-photolysis, we crystals exists state...