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T G Ebrey

Publications and source records attributed to T G Ebrey.

At least 73 records · Page 4Linked to original sources
PubMed1986

Resonance Raman study of the pink membrane photochemically prepared from the deionized blue membrane of H. halobium.

We report here the Resonance Raman spectrum of a 'pink' membrane (lambda max approximately 495 nm) photochemically generated from the deionized 'blue' membrane (Chang et al., 1985). Comparison of the Raman spectrum of the pink membrane with that of the model compounds, as well as the chromophore extraction data, indicate that the chromophore in the pink membrane is in the 9-cis configuration. The Schiff base peak at approximately 1,652 cm-1 shifts to approximately 1,622 cm-1 upon deuteration of the pink membrane, showing that the chromophore is bound to the bacterio-opsin by a protonated Schiff base linkage. The location of the Schiff base peak, as well as the 30 cm-1 shift that it undergoes upon deuteration, are quite different from the corresponding values for the native bacteriorhodopsin, suggesting differences in the local environment for the Schiff base in these pigments.

Halobacterium↗
PubMed1985

Fourier-transform infrared difference spectroscopy of rhodopsin and its photoproducts at low temperature.

Fourier-transform infrared difference spectroscopy has been used to detect the vibrational modes in the chromophore and protein that change in position or intensity between rhodopsin and the photoproducts formed at low temperature (70 K), bathorhodopsin and isorhodopsin. A method has been developed to obtain infrared difference spectra between rhodopsin and bathorhodopsin, bathorhodopsin and isorhodopsin, and rhodopsin and isorhodopsin. To aid in the identification of the vibrational modes, we performed experiments on deuterated and hydrated films of native rod outer segments and rod outer segments regenerated with either retinal containing 13C at carbon 15 or 15-deuterioretinal. Our infrared measurements provide independent verification of the resonance Raman result that the retinal in bathorhodopsin is distorted all-trans. The positions of the C = N stretch in the deuterated pigment and the deuterated pigments regenerated with 11-cis-15-deuterioretinal or 11-cis-retinal containing 13C at carbon 15 are indicative that the Schiff-base linkage is protonated in rhodopsin, bathorhodopsin, and isorhodopsin. Furthermore, the C = N stretching frequency occurs at the same position in all three species. The data indicate that the protonated Schiff base has a C = N trans conformation in all three species. Finally, we present evidence that, even in these early stages of the rhodopsin photosequence, changes are occurring in the opsin and perhaps the associated lipids.

Animals↗
PubMed1985

Characterization of the chromophore of the third rhodopsin-like pigment of Halobacterium halobium and its photoproduct.

Halobacterium halobium contains at least three retinal-containing pigments: bacteriorhodopsin, halorhodopsin, and a third rhodopsin-like pigment (tR) absorbing at approximately 590 nm, tR590. Illumination of tR590 gives rise to a very long-lived blue absorbing photoproduct, tR370. Using high-performance liquid chromatography we show that the chromophore of tR590 is primarily all-trans retinal and its conversion by light to tR370 causes the chromophore to isomerize primarily to the 13-cis conformation. Irradiation of the tR370 gives rise to a transient photoproduct absorbing at approximately 520 nm that decays back to the initial pigment tR590. In addition to all-trans retinal, the apomembrane of tR can also combine with 13-cis retinal but not with the 9- or 11-cis isomers.

Bacteriorhodopsins↗
PubMed1984

Resonance Raman study of the primary photochemistry of visual pigments. Hypsorhodopsin.

We report here the first resonance Raman results of octopus hypsorhodopsin, a species formed photochemically at very low temperatures from visual pigments. A pump-probe technique was used to obtain Raman spectra from samples at 12 degrees K whose photostationary state mixtures were either hypsorhodopsin rich or hysorhodopsin poor. The data strongly suggest that the Schiff-base linkage between the chromophore of hysorhodopsin and apoprotein is protonated. Further, the results suggest that hypsorhodopsin's chromophore is in some torsionally distorted conformation, possibly having torsional departures from an all-trans isomeric form.

Animals↗
PubMed1984

A correlation between proton pumping and the bacteriorhodopsin photocycle.

In an attempt to establish a relationship between proton pumping and the photocycle intermediates of bacteriorhodopsin, we have studied the effects of pH and temperature on flash-induced proton pumping and the photointermediates O640 and M412. The relative quantum yield of flash-induced proton pumping is both pH and temperature dependent. It is high in the acid pH range and at low temperatures but decreases in the basic pH range and at high temperatures. The decay of M412 is biphasic. The amplitude of the slowly decaying component (Ms) was found to be pH dependent with a pK similar to that of the delta pH. The pH dependence of the fast-decaying component (Mf) is opposite to that of Ms and delta pH. Like that of Ms, the amplitude of O640 is high in the acid pH range, but unlike the amplitude of Ms, it declines very rapidly at pHs greater than 6.5; the amplitude of O640 becomes zero around pH 8. The temperature dependence of the amplitude of Ms was found to be similar to that of delta pH, being high at low temperatures and low in the high-temperature range. The temperature dependence of Mf and O640 is opposite to that of delta pH and Ms. Whereas, O640 and Ms seem to be likely candidates (from the pH dependence) for coupling between proton pumping and the photocycle, Ms is more likely because (i) the relative quantum yield of both Ms and delta pH seem not to go to zero at higher pHs and (ii) Ms and delta pH have similar temperature dependence.

Bacteriorhodopsins↗
PubMed1983

Effects of pressure and temperature on the M412 intermediate of the bacteriorhodopsin photocycle. Implications for the phase transition of the purple membrane.

The effects of pressure and temperature on the decay kinetics of the M412 (M) intermediate in the photocycle of bacteriorhodopsin were studied to provide information about the phase transitions of the purple membrane lipids. The activation volume (delta V++) for the decay of M is expected to be different below and above a phase transition. However, no abrupt change in delta V++ was found from 3.5 degrees to 60 degrees C. But a sharp break was observed in a plot of the logarithm of the rate of M decay vs. pressure. Extrapolation of this break point to standard atmospheric pressure gives a temperature of -42 degrees C, which probably corresponds to the phase transition of the purple membrane lipids. This conclusion is supported by studies of the effect of pressure on the M kinetics of bacteriorhodopsin incorporated into dimyristoylphosphatidylcholine vesicles, whose phase transition has previously been characterized.

Bacteriorhodopsins↗
PubMed1983

Blue light effect on proton pumping by bacteriorhodopsin.

Proton pumping in closed vesicular systems containing bacteriorhodopsin that is initiated by an orange flash, is diminished by a subsequent blue flash. This blue light effect is due to light absorbed by the photocycle intermediate M412 (M), which was formed by the orange flash. A kinetic analysis of the blue-light-induced reduction of proton pumping shows that of the two components of M, only the slowly decaying component is involved in the reduction of proton movement. This may be the first correlation between a proton movement and a specific photochemical intermediate of bacteriorhodopsin. Furthermore, we report that blue light, acting on the slowly decaying intermediate, probably causes a movement of the protons in a direction opposite to that normally seen for light absorbed by bacteriorhodopsin.

Bacteriorhodopsins↗
PubMed1982

Effect of the removal of the COOH-terminal region of bacteriorhodopsin on its light-induced H+ changes.

Removal of the COOH-terminal region of bacteriorhodopsin by digestion with trypsin or papain reduces the yield of light-induced H+ release by 50-70%. The rate of H+ release is not affected significantly, but the half time of H+ uptake increases almost twofold. However, there is no effect on the photocycle of bacteriorhodopsin as judged by the yield and decay kinetics of the M412 photointermediate. The H+:M ratio in enzyme-digested membranes is approximately 0.4-0.8, whereas untreated membranes have a H+:M ratio of approximately 2. Purple membrane sheets stored in distilled water at 4 degrees C for prolonged periods also have a low H+:M ratio, probably due to protease activity associated with bacterial contamination. Electrophoresis on sodium dodecylsulfate-polyacrylamide gels showed that both the enzyme-treated and the stored purple membrane samples have a higher electrophoretic mobility compared to the fresh preparation. The reduction in molecular weight can be accounted for by the loss of several residues from the COOH-terminal portion of the bacteriorhodopsin. We propose that the COOH-terminal region is partially responsible for the high yield of H+ release by the purple membrane.

Bacteriorhodopsins↗
PubMed1981

Isolation of an inhibitory protein for the cyclic guanosine 3','5'-monophosphate phosphodiesterase of bovine rod outer segments.

Cyclic guanosine 3',5'-monophosphate phosphodiesterase in crude extracts from bovine rod outer segments can be activated by the addition of bleached rod outer segment membranes and GTP. In the absence of rhodopsin-containing membranes, the phosphodiesterase specific activity decreases with increasing concentration. A trypsin-sensitive inhibitor believed to be responsible for this phenomenon can be separated from the phosphodiesterase by DEAE-cellulose chromatography of the crude extract. Phosphodiesterase eluted from the DEAE-cellulose column shows considerably less concentration-dependence than in the crude extract. This partially purified phosphodiesterase was used as the substrate to assay for inhibitor. A GTPase which is active only in the presence of bleached rod outer segment membranes coelutes with the phosphodiesterase and is distinct from the phosphodiesterase inhibitor we have isolated.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed1981

Photochemical and functional properties of bacteriorhodopsins formed from 5,6-dihydro- and 5,6-dihydrodesmethylretinals.

5,6-Dihydroretinal and 5,6-dihydro-1,1,5,9,13-desmethylretinal are synthesized, and their all-trans isomers are shown to form pigment analogues (lambda max at 475 and 460 nm, respectively) of bacteriorhodopsin (purple membrane protein). The shift of the absorption maximum od the pigment from that of the protonated Schiff base of the chromophore for 5,6-dihydrobacteriorhodopsin is small compared to that of the native pigment, suggesting that negative charges similar to those controlling the lambda max of visual pigment rhodopsin exist near the cyclohexyl ring. Both pigment analogues undergo reversible light-induced spectral shifts reflecting cyclic photoreactions of the pigments. These results indicate that the absence of the C-5--C-6 double bond and of the five methyl groups of retinal does not abolish the photochemistry of these pigment analogues and strongly suggest that these structural features are not directly required for the photoreactions of native bacteriorhodopsin. The apparent rates of the photochemical transformations of these artificial pigments are quite different from those of bacteriorhodopsin. A working hypothesis is proposed for the photocycle of the pigment analogues, which includes a slower light-induced cycling rate (for the light-adapted pigments) than that of native bacteriorhodopsin and an increased rate of dark adaptation. When incorporated into egg lecithin vesicles both pigment analogues show proton pumping ability, again indicating that the missing double bond and the methyl groups are not structurally required for the function of the pigments.

Bacteriorhodopsins↗
PubMed1981

Flash photolysis and low temperature photochemistry of bovine rhodopsin with a fixed 11-ene.

Nonbleachable rhodopsins containing retinal moieties with fixed 11-ene structures have been prepared. When the nonbleachable rhodopsin analogue corresponding to the natural pigment was flash-photolysed at 20.8 degrees C, no absorption changes occurred at the monitoring wavelengths of 380, 480, and 580 nm for the time range of 2 microseconds--10 s. This observation is in contrast to that of natural rhodopsin which showed the formation of metarhodopsin I and its decay to meta II. Irradiation of the artificial rhodopsin, 77 K, with light of 460 and 540 nm, also gave no spectral changes; in the case of natural rhodopsin, however, the irradiation leads to formation of the red-shifted intermediate bathorhodopsin. The absence of photochemistry in the artificial pigment shows that an 11-cis to trans photoisomerization of the retinal moiety is a crucial step in inducing the chain of events in te photolysis of rhodopsin.

Animals↗
PubMed1981

Resonance Raman study of the primary photochemistry of bacteriorhodopsin.

Resonance Raman multicomponent spectra of the light-adapted form of bacteriorhodopsin, bRLA568, and its first photoproduct, K628, have been obtained at liquid nitrogen temperatures. The spectra of both bRLA568 and K628 could be obtained with the known sample compositions under our irradiating conditions and computer subtraction techniques. In agreement with previous results, we find that both bRLA568 and K628 contain chromophores linked to the apoprotein by protonated Schiff bases of retinal. Neither pigment form, suspended in H2O or 2H2O, compares closely to the spectral features of all-trans and 13-cis protonated and deuterated model chromophores, respectively. The data are consistent with other results, suggesting that a chromophore isomerization takes place in the bRLA568-to-K628 phototransition. However, the exact structure of the in situ chromophore would appear not to involve simple trans-to-13-cis structures found in solution.

Apoproteins↗