Impact of protein-chromophore interaction on the retinal excited state and photocycle of rhodopsin: role of conserved tryptophan residues.

The function of microbial as well as mammalian retinal proteins ( rhodopsins) is associated with a photocycle initiated by light excitation of the retinal chromophore of the protein, covalently bound through a protonated Schiff base linkage. Although electrostatics controls chemical reactions of many organic molecules, attempt to understand its role in controlling excited state reactivity of rhodopsins and, thereby, their photocycle is scarce. Here, we investigate the effect of highly conserved tryptophan residues, between which the all- retinal chromophore of the protein is sandwiched in microbial rhodopsins, on the charge distribution along the retinal excited state, quantum yield and nature of the light-induced photocycle and absorption properties of rhodopsin (GR).

Microsecond All-Optical Modulation by Biofunctionalized Porous Silicon Microcavity.

We successfully created a composite photonic structure out of porous silicon (PSi) microcavities doped by the photochromic protein, photoactive yellow protein (PYP). Massive incorporation of the protein molecules into the pores was substantiated by a 30 nm shift of the resonance dip upon functionalization, and light-induced reflectance changes of the device due to the protein photocycle were recorded. Model calculations for the photonic properties of the device were consistent with earlier results on the nonlinear optical properties of the protein, whose degree of incorporation into the PSi structure was also estimated.

Spectral and Redox Properties of a Recombinant Mouse Cytochrome 561 Protein Suggest Transmembrane Electron Transfer Function.

Cytochrome 561 proteins (CYB561s) are integral membrane proteins with six trans-membrane domains, two heme- redox centers, one on each side of the host membrane. The major characteristics of these proteins are their ascorbate reducibility and trans-membrane electron transferring capability. More than one CYB561 can be found in a wide range of animal and plant phyla and they are localized in membranes different from the membranes participating in bioenergization.

Kinetics and Energetics of Intramolecular Electron Transfer in Single-Point Labeled TUPS-Cytochrome Derivatives.

Electron transfer within and between proteins is a fundamental biological phenomenon, in which efficiency depends on several physical parameters. We have engineered a number of horse heart cytochrome single-point mutants with cysteine substitutions at various positions of the protein surface. To these cysteines, as well as to several native lysine side chains, the photoinduced redox label 8-thiouredopyrene-1,3,6-trisulfonate (TUPS) was covalently attached.

Optically Manipulated Microtools to Measure Adhesion of the Nanoparticle-Targeting Ligand Glutathione to Brain Endothelial Cells.

Targeting nanoparticles as drug delivery platforms is crucial to facilitate their cellular entry. Docking of nanoparticles by targeting ligands on cell membranes is the first step for the initiation of cellular uptake. As a model system, we studied brain microvascular endothelial cells, which form the anatomical basis of the blood-brain barrier, and the tripeptide glutathione, one of the most effective targeting ligands of nanoparticles to cross the blood-brain barrier.

Machine-learning model selection and parameter estimation from kinetic data of complex first-order reaction systems.

Dealing with a system of first-order reactions is a recurrent issue in chemometrics, especially in the analysis of data obtained by spectroscopic methods applied on complex biological systems. We argue that global multiexponential fitting, the still common way to solve such problems, has serious weaknesses compared to contemporary methods of sparse modeling. Combining the advantages of group lasso and elastic net-the statistical methods proven to be very powerful in other areas-we created an optimization problem tunable from very sparse to very dense distribution over a large pre-defined grid of time constants, fitting both simulated and experimental multiwavelength spectroscopic data with high computational efficiency.

Differential Polarization Imaging of Plant Cells. Mapping the Anisotropy of Cell Walls and Chloroplasts.

Modern light microscopy imaging techniques have substantially advanced our knowledge about the ultrastructure of plant cells and their organelles. Laser-scanning microscopy and digital light microscopy imaging techniques, in general-in addition to their high sensitivity, fast data acquisition, and great versatility of 2D-4D image analyses-also opened the technical possibilities to combine microscopy imaging with spectroscopic measurements. In this review, we focus our attention on differential polarization (DP) imaging techniques and on their applications on plant cell walls and chloroplasts, and show how these techniques provided unique and quantitative information on the anisotropic molecular organization of plant cell constituents: (i) We briefly describe how laser-scanning microscopes (LSMs) and the enhanced-resolution Re-scan Confocal Microscope (RCM of Confocal.

Nonlinear Optical Investigation of Microbial Chromoproteins.

Membrane-bound or cytosolic light-sensitive proteins, playing a crucial role in energy- and signal-transduction processes of various photosynthetic microorganisms, have been optimized for sensing or harvesting light by myriads of years of evolution. Upon absorption of a photon, they undergo a usually cyclic reaction series of conformations, and the accompanying spectro-kinetic events assign robust nonlinear optical (NLO) properties for these chromoproteins. During recent years, they have attracted a considerable interest among researchers of the applied optics community as well, where finding the appropriate NLO material for a particular application is a pivotal task.

Determination of the p Values of -Resveratrol, a Triphenolic Stilbene, by Singular Value Decomposition. Comparison with Theory.

Several independent determinations of the p values of -resveratrol in water have led to conflicting results. Singular value decomposition analysis of UV absorption spectra of -resveratrol (Resv) in N-outgased aqueous solutions buffered to pH values in the 7.0-13.

Accumulation of 2-Acetylamino-5-mercapto-1,3,4-thiadiazole in chitosan coatings for improved anticorrosive effect on zinc.

Chitosan (Chit) coatings were applied on zinc substrates by the dip-coating method. Subsequently, the coatings were impregnated with a corrosion inhibitor, 2-Acetylamino-5-mercapto-1,3,4-thiadiazole (AcAMT) to obtain an increased anticorrosive effect. The coating thickness and the AcAMT accumulation were determined using UV-Vis spectroscopy on glass and quartz substrates, respectively.

Translational Diffusion and Unstable Conformer Trapping in Glassy Isopentane at 77 K.

Diffusional quenching in isopentane (IP) glass at 77 K is demonstrated by the reduction of triphenylene phosphorescence lifetimes in the presence of 1,3-pentadiene and/or molecular oxygen. Fluorescence spectra and lifetimes of - and -1,2-di(1-methyl-2-naphthyl)ethene in IP glass at 77 K reveal that the → photoisomerization leads to the trapping of unstable conformers of the isomer. The claim that IP at 77 K is not sufficiently viscous to trap unstable photoproduct conformers is invalidated.

Lidocaine turns the surface charge of biological membranes more positive and changes the permeability of blood-brain barrier culture models.

The surface charge of brain endothelial cells forming the blood-brain barrier (BBB) is highly negative due to phospholipids in the plasma membrane and the glycocalyx. This negative charge is an important element of the defense systems of the BBB. Lidocaine, a cationic and lipophilic molecule which has anaesthetic and antiarrhytmic properties, exerts its actions by interacting with lipid membranes.

Fluorescence-detected linear dichroism imaging in a re-scan confocal microscope equipped with differential polarization attachment.

Confocal laser scanning microscopy is probably the most widely used and one of the most powerful techniques in basic biology, medicine and material sciences that is employed to elucidate the architecture of complex cellular structures and molecular macro-assemblies. It has recently been shown that the information content, signal-to-noise ratio and resolution of such microscopes (LSMs) can be improved significantly by adding different attachments or modifying their design, while retaining their user-friendly features and relatively moderate costs. Differential polarization (DP) attachments, using high-frequency modulation/demodulation circuits, have made LSMs capable of high-precision 2D and 3D mapping of the anisotropy of microscopic samples-without interfering with their 'conventional' fluorescence or transmission imaging (Steinbach et al.

Spectrokinetic characterization of photoactive yellow protein films for integrated optical applications.

In this paper, the photocycle of the dried photoactive yellow protein film has been investigated in different humidity environments, in order to characterize its nonlinear optical properties for possible integrated optical applications. The light-induced spectral changes of the protein films were monitored by an optical multichannel analyser set-up, while the accompanying refractive index changes were measured with the optical waveguide lightmode spectroscopy method. To determine the number and kinetics of spectral intermediates in the photocycle, the absorption kinetic data were analysed by singular value decomposition and multiexponential fitting methods, whose results were used in a subsequent step of fitting a photocycle model to the data.

Interactions at the Peptide/Silicon Surfaces: Evidence of Peptide Multilayer Assembly.

Selective deposition of peptides from liquid solutions to n- and p-doped silicon has been demonstrated. The selectivity is governed by peptide/silicon adhesion differences. A noninvasive, fast characterization of the obtained peptide layers is required to promote their application for interfacing silicon-based devices with biological material.

The trans-membrane cytochrome b561 proteins: structural information and biological function.

Cytochrome b561 (CYB561) proteins are ascorbate reducible, trans-membrane proteins consisting of 200-300 amino acids, about half of which are hydrophobic. The first identified CYB561 protein was discovered more than 40 years ago, and is localized in the chromaffin granule membrane of the mammalian adrenal glands. Proteins with similar structural elements and biophysical and biochemical properties were identified in a wide range of animal and plant phyla in the past 15 years.

Disentangling electron tunneling and protein dynamics of cytochrome c through a rationally designed surface mutation.

Nonexponential distance dependence of the apparent electron-transfer (ET) rate has been reported for a variety of redox proteins immobilized on biocompatible electrodes, thus posing a physicochemical challenge of possible physiological relevance. We have recently proposed that this behavior may arise not only from the structural and dynamical complexity of the redox proteins but also from their interplay with strong electric fields present in the experimental setups and in vivo (J. Am Chem.

Effect of Hofmeister cosolutes on the photocycle of photoactive yellow protein at moderately alkaline pH.

The photocycle of photoactive yellow protein was studied by kinetic absorption spectroscopy from below 100ns to seconds, at moderately alkaline pH, in the presence of high concentrations of various salts. Chemometric analysis combined with multiexponential fit of the flash-induced difference spectra provided evidence for five intermediates, including a spectrally silent form before the final recovery of the parent state, but only three with significantly distinct spectra. The calculated intermediate spectra constituted the input for the following spectrotemporal model fit using a sufficiently complex photocycle scheme with reversible transitions.

Root canal hydrophobization by dentinal silanization: improvement of silicon-based endodontic treatment tightness.

A new strategy to improve silicon-based endodontic treatment tightness by dentine hydrophobization is presented in this work: root dentine was silanized to obtain a hydrophobic dentine-sealer interface that limits fluid penetration. This strategy was based on the grafting of aliphatic carbon chains on the dentine through a silanization with the silane end groups [octadecyltrichlorosilane (OTS) and octadecyltriethoxysilane]. Dentine surface was previously pretreated, applying ethylenediaminetetraacetic acid and sodium hypochlorite, to expose hydroxyl groups of collagen for the silane grafting.

Porous silicon/photosynthetic reaction center hybrid nanostructure.

The purified photosynthetic reaction center protein (RC) from Rhodobacter sphaeroides R-26 purple bacteria was bound to porous silicon microcavities (PSiMc) either through silane-glutaraldehyde (GTA) chemistry or via a noncovalent peptide cross-linker. The characteristic resonance mode in the microcavity reflectivity spectrum red shifted by several nanometers upon RC binding, indicating the protein infiltration into the porous silicon (PSi) photonic structure. Flash photolysis experiments confirmed the photochemical activity of RC after its binding to the solid substrate.

Light-harvesting bio-nanomaterial using porous silicon and photosynthetic reaction center.

Porous silicon microcavity (PSiMc) structures were used to immobilize the photosynthetic reaction center (RC) purified from the purple bacterium Rhodobacter sphaeroides R-26. Two different binding methods were compared by specular reflectance measurements. Structural characterization of PSiMc was performed by scanning electron microscopy and atomic force microscopy.

Dihydrolipoic acid reduces cytochrome b561 proteins.

Cytochrome b561 (Cyt-b561) proteins constitute a family of trans-membrane proteins that are present in a wide variety of organisms. Two of their characteristic properties are the reducibility by ascorbate (ASC) and the presence of two distinct b-type hemes localized on two opposite sides of the membrane. Here we show that the tonoplast-localized and the putative tumor suppressor Cyt-b561 proteins can be reduced by other reductants than ASC and dithionite.

Mapping local electric fields in proteins at biomimetic interfaces.

We present a novel approach for determining the strength of the electric field experienced by proteins immobilised on membrane models. It is based on the vibrational Stark effect of a nitrile label introduced at different positions on engineered proteins and monitored by surface enhanced infrared absorption spectroscopy.