Biomedical subjects

L Keszthelyi

Publications and source records attributed to L Keszthelyi.

At least 19 recordsLinked to original sources

PubMed2006

Excitation of the L intermediate of bacteriorhodopsin: electric responses to test x-ray structures.

The L intermediate of bacteriorhodopsin was excited, and its electrical response was measured. Two positive components were found in it with respect to the direction of proton pumping: an unresolved fast component, and a slower one (tau=7 micros) of small amplitude. The fast component was assigned to a charge motion corresponding to reisomerization of the retinal moiety, whereas the slow one was attributed to charge rearrangements reestablishing the ground state. Because three x-ray crystallographic structures have recently been reported for the L intermediate, it seemed important to calculate the intramolecular dipole moment changes associated to bR-->L for all three structures, so as to compare them with similar quantities determined from the electrical signals. The results are discussed in terms of amino acid side chains possibly contributing to the observed effect. We propose to use electrical signals as a verification tool for intermediate structures of the photocycle, and thus for molecular models of proton pumping.

Bacteriorhodopsins↗

PubMed2005

Actinic light-energy dependence of proton release from bacteriorhodopsin.

Measuring the light-density (fluence) dependence of proton release from flash excited bacteriorhodopsin with two independent methods we found that the lifetime of proton release increases and the proton pumping activity, defined as a number of protons per number of photocycle, decreases with increasing fluence. An interpretation of these results, based on bending of purple membrane and electrical interaction among the proton release groups of bacteriorhodopsin trimer, is presented.

Bacteriorhodopsins↗

PubMed2003

Late events in the photocycle of bacteriorhodopsin mutant L93A.

In the photocycle of bacteriorhodopsin (bR) from Halobacterium salinarum mutant L93A, the O-intermediate accumulates and the cycling time is increased approximately 200 times. Nevertheless, under continuous illumination, the protein pumps protons at near wild-type rates. We excited the mutant L93A in purple membrane with single or triple laser flashes and quasicontinuous illumination, (i.e., light for a few seconds) and recorded proton release and uptake, electric signals, and absorbance changes. We found long-living, correlated, kinetic components in all three measurements, which-with exception of the absorbance changes-had not been seen in earlier investigations. At room temperature, the O-intermediate decays to bR in two transitions with rate constants of 350 and 1800 ms. Proton uptake from the cytoplasmic surface continues with similar kinetics until the bR state is reestablished. An analysis of the data from quasicontinuous illumination and multiple flash excitation led to the conclusion that acceleration of the photocycle in continuous light is due to excitation of the N-component in the fast N<-->O equilibrium, which is established at the beginning of the severe cycle slowdown. This conclusion was confirmed by an action spectrum.

Adaptation, Physiological↗

PubMed2003

Photoexcitation of the O-intermediate in bacteriorhodopsin mutant L93A.

During the extended lifetime of the O-state in bacteriorhodopsin (bR) mutant L93A, two substates have been distinguished. The first O-intermediate (OI) is in rapid equilibrium with N and apparently still has a 13-cis chromophore. OI undergoes a photoreaction with a small absorbance change, positive charge transport in the pumping direction, and proton release and uptake. None of these effects was detected after photoexcitation of the late O (OII). The most likely interpretation of the effects seen is an accelerated return of the molecule from the OI- to the bR-state. However, with a lifetime approximately 140 ms, the reaction cannot account for the observed high pumping efficiency of the mutant under continuous illumination. We suggest that OII corresponds to the O-intermediate with a twisted all-trans chromophore seen in the photocycle of wild-type bR, where the 13-cis OI-intermediate under the usual conditions does not accumulate in easily detectable amounts and, therefore, has generally been overlooked. Both the OI- and OII-decays are apparently strongly inhibited in the mutant.

Adaptation, Physiological↗

PubMed2001

Electric signals of light excited bacteriorhodopsin mutant D96N.

The study of mutant D96N played an important role in understanding proton translocation by light driven bacteriorhodopsin. Our measurement of photoelectric current for single and double flash illumination revealed new details of the photocycle of this mutant. With double flash excitation we found an intermediate absorbing near the wavelength of the ground state of bacteriorhodopsin (bR) but pumping in the opposite direction. This intermediate has the same lifetime as the species described by Zimányi et al. [Proc. Natl. Acad. Sci. USA 96 (1999) 4414-4419] and was assigned to early recovery of a fraction of the ground state after excitation. Because the electric response does not reconcile with that of the ground state, we tentatively assign it to the L intermediate or to an intermediate similar in absorption to bR (bR').

Bacteriorhodopsins↗

PubMed2001

Buffer effects on electric signals of light-excited bacteriorhodopsin mutants.

The effects of glycyl-glycine and bis-trispropane buffers on the light-excited electric signals due to proton motion in the molecule were studied for the bacteriorhodopsin (bR) mutants D38R, D96N, E204Q, R227Q, D85N, D85T, R82Q/D85N, and D85N/D96N in purple membranes and for delipidated purple membrane containing the wild-type bR. The results show additional charge motion caused by the buffers in all cases. Arrhenius parameters calculated from the temperature dependence of the difference signals (with buffer minus without buffer) are similar to the parameters found for the wild-type bR in the case of these buffers: the values of the activation enthalpies are mostly in the range 25-50 kJ/mol; all the activation entropies are negative. The results are evaluated with the cluster hypothesis outlined previously.

Amino Acid Substitution↗

PubMed2001

Proton binding to biological membranes.

Biological membranes contain proton-binding moieties. A laser-induced proton pulse was used to characterize the proton-binding properties of bacterioopsin-containing membranes and of sarcoplasmic reticulum. Different protonation and deprotonation processes occurred. The liberation of protons from pyranine dye and the protonation of the membranes were independent of temperature; the reprotonation of pyranine and proton release from the membranes were temperature dependent. In the cases of membrane-free and membrane-containing systems, the activation enthalpies and entropies were calculated from the decay rates. The activation enthalpy of 16 kJ/mol for reprotonation of pyranine in membrane-free solution is characteristic for a diffusion-controlled process. The value for the membrane-containing systems was nearly double, suggesting that the buffering moieties of the membrane surfaces strongly bind the protons, raising the activation enthalpies. This is possibly an effect of the Coulomb cages formed from closely located proton acceptor sites. The activation entropies were positive in all cases.

Arylsulfonates↗

PubMed2001

Homochirality of biomolecules: counter-arguments against critical notes.

Three recently published critical papers by Bonner and coworkers on the extraterrestrial origin of the homochirality of biomolecules and the amplification of tiny enantiomeric excess are discussed. The presented arguments show the difficulties involved in circularly polarized u.v. and visible synchrotron radiation from neutron stars, in their ratio to non circularly polarized light and in racemization of the products. Attention is called upon another mechanism for production of extraterrestrial handedness based on Salam's condensation theory and on the recent experimental demonstration of the enantioselective magnetochiral photochemistry. Arguments, as well as experiments, point out that tiny enantiomeric excesses might be amplified via the Yamagata accumulation principle questioned by Bonner.

Extraterrestrial Environment↗

PubMed2001

Charge Motion during the Photocycle of Bacteriorhodopsin.

The function of bacteriorhodopsin in Halobacterium salinarum is to pump protons from the internal side of the plasma membrane to the external after light excitation, thereby building up electrochemical energy. This energy is transduced into biological energy forms. This review deals with one of the methods elaborated for recording the charge transfer inside the protein. In this method the current produced in oriented purple membrane containing bacteriorhodopsin is measured. It is shown that this method might be applied not only to correlate charge motion with the photocycle reactions but also for general problems like effect of water, electric field, and different ions and buffers for the functioning of proteins.

Bacteriorhodopsins↗

PubMed2000

Galileo at Io: results from high-resolution imaging.

During late 1999/early 2000, the solid state imaging experiment on the Galileo spacecraft returned more than 100 high-resolution (5 to 500 meters per pixel) images of volcanically active Io. We observed an active lava lake, an active curtain of lava, active lava flows, calderas, mountains, plateaus, and plains. Several of the sulfur dioxide-rich plumes are erupting from distal flows, rather than from the source of silicate lava (caldera or fissure, often with red pyroclastic deposits). Most of the active flows in equatorial regions are being emplaced slowly beneath insulated crust, but rapidly emplaced channelized flows are also found at all latitudes. There is no evidence for high-viscosity lava, but some bright flows may consist of sulfur rather than mafic silicates. The mountains, plateaus, and calderas are strongly influenced by tectonics and gravitational collapse. Sapping channels and scarps suggest that many portions of the upper approximately 1 kilometer are rich in volatiles.

Extraterrestrial Environment↗

PubMed2000

A close-up look at Io from Galileo's near-infrared mapping spectrometer.

Infrared spectral images of Jupiter's volcanic moon Io, acquired during the October and November 1999 and February 2000 flybys of the Galileo spacecraft, were used to study the thermal structure and sulfur dioxide distribution of active volcanoes. Loki Patera, the solar system's most powerful known volcano, exhibits large expanses of dark, cooling lava on its caldera floor. Prometheus, the site of long-lived plume activity, has two major areas of thermal emission, which support ideas of plume migration. Sulfur dioxide deposits were mapped at local scales and show a more complex relationship to surface colors than previously thought, indicating the presence of other sulfur compounds.

Extraterrestrial Environment↗

PubMed2000

Prometheus: Io's wandering plume.

Unlike any volcanic behavior ever observed on Earth, the plume from Prometheus on Io has wandered 75 to 95 kilometers west over the last 20 years since it was first discovered by Voyager and more recently observed by Galileo. Despite the source motion, the geometric and optical properties of the plume have remained constant. We propose that this can be explained by vaporization of a sulfur dioxide and/or sulfur "snowfield" over which a lava flow is moving. Eruption of a boundary-layer slurry through a rootless conduit with sonic conditions at the intake of the melted snow can account for the constancy of plume properties.

Cold Temperature↗

PubMed2000

Buffer effects on electric signals of light-excited bacteriorhodopsin.

Buffers change the electric signals of light-excited bacteriorhodopsin molecules in purple membrane if their concentration and the pH of the low-salt solution are properly selected. "Positive" buffers produce a positive component, and "negative" buffers a negative component in addition to the signals due to proton pumping. Measurement of the buffer effects in the presence of glycyl-glycine or bis-tris propane revealed an increase of approximately 2 and a change of sign and a decrease to approximately -0.5 in the translocated charge in these cases, respectively. These factors do not depend on temperature. The Arrhenius parameters established from the evaluation of the kinetics indicate activation enthalpies of 35-40 kJ/mol and negative activation entropies for the additional signals. These values agree with those found by surface-bound pH-sensitive probes in the search of the timing of proton release and uptake. The electric signals were also measured in the case of D(2)O solutions with similar results, except for the increased lifetimes. We offer a unified explanation for the data obtained with surface-bound probes and electric signals based on the clusters at extracellular and cytoplasmic sites of bacteriorhodopsin participating in proton release and uptake.

Bacteriorhodopsins↗

PubMed1999

Photoelectric response of the N intermediate of bacteriorhodopsin and its mutant T46V.

Double flash experiments were performed in order to gain information about the characteristics of the N intermediates of the photocycle of bacteriorhodopsin. The N intermediates of wild-type bacteriorhodopsin and mutant T46V were excited at different delay times after the first laser flash which induced the photocycle and the electric responses were registered. These electric signals revealed that charge motions occurred in both cases, though charge translocation, i.e. H(+) pumping, could not be observed. The delay time dependence of the electric signals is characterized by two distinct processes corresponding to two substates of the N intermediates.

Bacteriorhodopsins↗

PubMed1999

Demonstration of the parity-violating energy difference between enantiomers.

Racemic mixtures of (+) and (-) sodium potassium tartrate, tris(1, 2-ethanediamine)cobalt(III), and tris(1,2-ethanediamine)iridium(III) molecules were crystallized, and the optical activities of the resulting crystalline materials, dissolved in water, were carefully measured to study the influence of the parity-violating energy difference in the crystallization process. Although no effect was found in the case of tartrate, enantiomeric excess appeared in the crystallization of the cobalt and iridium complexes. These investigations, performed in our laboratory, demonstrated the contribution of the parity-violating neutral weak current to the forces acting in molecules.

Circular Dichroism↗

PubMed1999

Interpretation of the spatial charge displacements in bacteriorhodopsin in terms of structural changes during the photocycle.

We have recently introduced a method, made possible by an improved orienting technique using a combination of electric and magnetic fields, that allows the three-dimensional detection of the intramolecular charge displacements during the photocycle of bacteriorhodopsin. This method generates electric asymmetry, a prerequisite for the detection of electric signal on the macroscopic sample, in all three spatial dimensions. Purple membrane fragments containing bacteriorhodopsin were oriented so that their permanent electric dipole moment vectors were perpendicular to the membrane plane and pointed in the same direction. The resulting cylindrical symmetry was broken by photoselection, i. e., by flash excitation with low intensity linearly polarized light. From the measured electric signals, the three-dimensional motion of the electric charge center in the bacteriorhodopsin molecules was calculated for the first 400 microseconds. Simultaneous absorption kinetic recording provided the time-dependent concentrations of the intermediates. Combining the two sets of data, we determined the discrete dipole moments of intermediates up to M. When compared with the results of current molecular dynamics calculations, the data provided a decisive experimental test for selecting the optimal theoretical model for the proton transport and should eventually lead to a full description of the mechanism of the bacteriorhodopsin proton pump.

Bacteriorhodopsins↗

PubMed1998

High-temperature silicate volcanism on Jupiter's moon Io.

Infrared wavelength observations of Io by the Galileo spacecraft show that at least 12 different vents are erupting lavas that are probably hotter than the highest temperature basaltic eruptions on Earth today. In at least one case, the eruption near Pillan Patera, two independent instruments on Galileo show that the lava temperature must have exceeded 1700 kelvin and may have reached 2000 kelvin. The most likely explanation is that these lavas are ultramafic (magnesium-rich) silicates, and this idea is supported by the tentative identification of magnesium-rich orthopyroxene in lava flows associated with these high-temperature hot spots.

Extraterrestrial Environment↗

PubMed1997

N-like intermediate in the photocycle of the acid purple form of bacteriorhodopsin.

The photocycle of the acid-purple form of bacteriorhodopsin (BR) was studied by multiwavelength absorption kinetic measurements. The data were evaluated using a new method based on the differences in the kinetics of the absorption changes at different wavelengths [Tokaji, Zs. (1995) FEBS Lett. 357, 156-160]. The evaluation indicates that, in contrast to the previous suggestion, the photocycle of the acid purple BR contains an intermediate that, in its spectral and kinetic properties, resembles the N intermediate of the alkaline BR photocycle.

Bacteriorhodopsins↗