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R M Hochstrasser

Publications and source records attributed to R M Hochstrasser.

35 records · Page 2Linked to original sources
PubMed1992

Picosecond dynamics of bacteriorhodopsin, probed by time-resolved infrared spectroscopy.

The photoinduced reaction cycle of bacteriorhodopsin (BR) has been studied by means of a recently developed picosecond infrared spectroscopic method at ambient temperature. BR - K difference spectra between 1560 and 1700 cm-1 have been recorded at delay times from 100 ps to 14 ns. The spectrum remains unchanged during this period. The negative difference OD band at 1660 cm-1 indicates the peptide backbone responds within 50 ps. A survey in the region of carboxylic side chain absorption around 1740 cm-1 reveals that perturbations of those groups, present in low-temperature FTIR spectra, are not observable within 10 ns, suggesting a slow conformational change.

Bacteriorhodopsins↗
PubMed1991

Ultrafast infrared spectroscopy of bacteriorhodopsin.

Picosecond infrared spectroscopy is developed and used for the first time to study the dynamics of photoexcited bacteriorhodopsin (BR). Both spectral and time-resolved data are obtained. The results open an entirely new approach to investigations of the BR photocycle. The infrared difference spectrum (K minus BR570) recorded at ambient temperature between 1,560 and 1,700 cm-1 is not identical with the spectrum reported for a frozen sample. Three bands of the K state at 1,622, 1,610, and 1,580 cm-1 and the bleaching at 1,637 cm-1 (C = NH stretch) are seen. These new spectral lines appear in less than 10 ps.

Bacteriorhodopsins↗
PubMed1989

Direct observations of ligand dynamics in hemoglobin by subpicosecond infrared spectroscopy.

The photodissociation of CO from HbCO at ambient temperature is studied by means of a femtosecond IR technique. The bleaching of the FeCO absorption and the appearance of a new IR absorption near that of free CO are both observed at 300 fs after optical excitation. The bleach does not recover on the time scale of a few picoseconds but does recover by approximately 4% within 1 ns, which suggests that a barrier to recombination is formed within a few picoseconds. The CO spectrum does not change significantly between 300 fs and 1 ns, suggesting that the CO quickly finds some locations in the heme pocket that are not more than a few angstroms from the iron. The de-ligated CO appears in its ground vibrational level. There is evidence that 85 +/- 10% of this CO remains in the heme pocket at 1 ns; it probably resides there for 50 ns. The flow of excess vibrational energy from the heme to the solvent was directly observed in the IR experiments. The heme cools within 1-2 ps while thermal disruption of the surrounding solvent structure requires approximately 30 ps.

Carboxyhemoglobin↗
PubMed1988

Picosecond transient absorption study of photodissociated carboxy hemoglobin and myoglobin.

The optical transient absorption spectra at 30 ps and 6.5 ns after photolysis are compared for both carboxy hemoglobin (HbCO) and carboxy myoglobin (MbCO). Both 355- and 532-nm excitation pulses were used. In all cases the shapes of the optical difference spectra thus generated are stationary over the complete time-scale studied. The photolysis spectra for MbCO are not significantly different from the equilibrium difference spectra generated on the same picosecond spectrometer when measured to an accuracy of +/- 0.5 nm. In addition, spectral parameters for delegated HbCO generated on the same spectrometer but detected by two different techniques, either by a Vidicon detector or point by point with photomultiplier tubes, are reported; the results are different from some of the previously reported picosecond experiments.

Animals↗
PubMed1988

Iron-carbonyl bond geometries of carboxymyoglobin and carboxyhemoglobin in solution determined by picosecond time-resolved infrared spectroscopy.

The iron-carbonyl geometries in carboxymyoglobin (MbCO) and carboxyhemoglobin (HbCO) in ambient temperature solution have been investigated using picosecond time-resolved infrared spectroscopy. Polarized infrared and visible beams were used to monitor the change in infrared absorbance of the bound CO stretch bands on photodissociation of the ligand. The ratio of the change in absorbance for perpendicular and parallel relative polarizations of the photolysis and infrared probe beams is directly related to the angle between the ligand bond axis and the normal to the heme plane. Ratios, and hence the angles, have been obtained for the configurations giving rise to the principal CO stretch infrared absorption bands of HbCO and MbCO: 18 degrees for the 1951 cm-1 band of HbCO; 20 degrees and 35 degrees, respectively, for the 1944 cm-1 and 1933 cm-1 bands of MbCO. Structures consistent with x-ray diffraction and the picosecond experiments reported here are proposed for MbCO and HbCO in which the Fe-C bond tilts to the heme normal and the Fe-C-O angle differs significantly from 180 degrees.

Carbon Monoxide↗
PubMed1987

Fluorescence and energy transfer of tryptophans in Aplysia myoglobin.

The fluorescence decay of tryptophan residues in apo and met Aplysia limacina myoglobin and sperm whale myoglobin were measured in aqueous solution at 10 degrees-15 degrees C. In all species, multiexponential behavior was observed in which the individual components displayed unique frequency-dependent emission characteristics. The results suggest that the tryptophan fluorescence in all met samples are quenched by rapid Forster energy transfer to the heme as predicted from the crystal geometry. Fluorescence from the apo protein is similar to that in solutions of free tryptophans. In addition, the fluorescence properties of the reversible thermal denaturation of Aplysia limacina met myoglobin was investigated between 25 degrees and 75 degrees C.

Animals↗
PubMed1987

Molecular dynamics simulations of fluorescence polarization of tryptophans in myoglobin.

The fluorescence of heme proteins is influenced by energy transfer from the excited tryptophan to the heme. Molecular dynamics simulations of the tryptophan and heme motions in sperm whale myoglobin were used to calculate the fluorescence intensity and anisotropy decays. The side chains underwent both small rapid orientational fluctuations and large infrequent transitions between conformations. The predicted motions of the tryptophans and the heme produce large fluctuations in the instantaneous rate of energy transfer, but no stable conformations in which energy transfer is suppressed were found. The calculated fluorescence anisotropies exhibited a large subpicosecond decay, corresponding to nondiffusive side-chain motions. The calculations adequately predict the observed fluorescence decay curve for myoglobin and the total anisotropy decay at 16-ps time resolution. The subnanosecond decays of anisotropy for tryptophan-14 in tuna myoglobin are not reproduced by the calculation.

Animals↗
PubMed1986

Molecular dynamics simulations of cooling in laser-excited heme proteins.

In transient optical experiments the absorbed photon raises the vibrational temperature of the chromophore. In heme proteins at room temperature conversion of a 530-nm photon into vibrational energy is estimated to raise the temperature of the heme by 500-700 K. Cooling of the heme is expected to occur mainly by interacting with the surrounding protein. We report molecular dynamics simulations for myoglobin and cytochrome c in vacuo that predict that this cooling occurs on the ps time scale. The decay of the vibrational temperature is nonexponential with about 50% loss occurring in 1-4 ps and with the remainder in 20-40 ps. These results predict the presence of nonequilibrium vibrational populations that would introduce ambiguity into the interpretation of transient ps absorption and Raman spectra and influence the kinetics of sub-ns geminate recombination.

Biophysical Phenomena↗
PubMed1985

Picosecond resonance Raman evidence for unrelaxed heme in the (carbonmonoxy)myoglobin photoproduct.

An actively and passively mode-locked Nd:YAG laser, producing 30-ps pulses of 1-mJ energy at 532 nm, has been used to photolyze (carbonmonoxy)myoglobin (MbCO) and generate its resonance Raman spectrum, which was recorded with a vidicon multichannel analyzer. The photoproduct spectrum was obtained by subtraction of the MbCO spectrum, obtained at lower incident power levels. Comparison with the spectrum of deoxyMb, obtained with the same apparatus, revealed frequency downshifts of approximately 4 cm-1, for bands at 1604, 1554, and 1542 cm-1, which are identified with porphyrin skeletal modes v10, v19, and v11. These frequencies are known to correlate inversely with the core size of the porphyrin ring, and the shifts imply a larger core size for the photoproduct than for deoxyMb. Similar shifts have been observed for the (carbonmonoxy)hemoglobin (HbCO) photoproduct; in that case, the shifts persist for longer than 20 ns, whereas they are absent in the MbCO photoproduct spectrum within 7 ns of photolysis. The unrelaxed state of the heme group region is therefore suggested to be maintained by protein forces, which relax more rapidly for Mb than Hb. This may reflect a tighter coupling in Hb of the out-of-plane movement of the Fe atom with the proximal histidine-containing F helix.

Animals↗
PubMed1984

Picosecond fluorescence decay of tryptophans in myoglobin.

The fluorescence decay characteristics of Mb, MbCO, metMb (sperm whale), metMb (yellowfin tuna), and their apo derivatives were determined by using a picosecond streak camera and time-correlated single photon counting. The emission is dominated by tryptophans that transfer their energy to the heme on a subnanosecond time scale. Sperm whale Mb and derivatives have two tryptophans and their decays can be interpreted mainly as two exponentials, one of ca. 20 ps and the other of 130 ps, whereas tuna Mb has one tryptophan and its emission is nonexponential but dominated by one component of 31 ps. These results along with Förster energy transfer calculations allow us to assign the ca. 30-ps emission to Trp-14 and the 130-ps emission to Trp-7 in Mb. The streak camera was modified to determine the decay of the fluorescence anisotropy. In metMb (tuna) the fluorescence anisotropy decays in 100 ps, which is postulated to result from rapid motion of the Trp-14. Because energy transfer was used to gate the anisotropy, the fast motion of Trp-14 is proposed to correspond to only 10% of the equilibrium distribution of molecules.

Animals↗
PubMed1983

Ultrafast relaxation in picosecond photolysis of nitrosylhemoglobin.

We report the successful observation of a picosecond transient difference spectrum in human nitrosylhemoglobin. The sample (23 degrees C) is excited with an approximately 8 ps, approximately 10 microJ pulse at 353 nm that generates a prompt transient having a two-component decay: the first is approximately exponential with tau = 17 +/- 4 ps, whereas the second is much weaker with an approximate tau = 100 ps. At slightly lower temperature (4 degrees C), the spectrum and time dependence are essentially unchanged. In contrast to our previous observations on carboxyhemoglobin and oxyhemoglobin, we find no long-lived photoproduct in nitrosylhemoglobin. We tentatively attribute the 17 +/- 4 ps decay to geminate recombination. These results, in conjunction with our previous work in HbO2 and HbCO, show that the rate of geminate recombination for 5 ns greater than tau greater than 5 ps increases through the series HbCO less than HbO2 less than HbNO. We note that trends are also seen for microsecond recombination rates HbCO less than HbO2 approximately equal to HbNO and for the kinetic co-operativity ratio HbCO greater than HbO2 greater than HbNO. A "critical on-barrier" model is presented that provides a consistent representation of these results. We suggest that spin-orbit effects could be a major contribution to the different recombination characteristics exhibited by the three ligands.

Carboxyhemoglobin↗
PubMed1981

Different dissociation pathways and observation of an excited deoxy state in picosecond photolysis of oxy- and carboxymyoglobin.

Picosecond transient absorption spectra of Mb, MbCO, and MbO2 have been studied at time delays of up to 10 ns after excitation at 353 nm. Particular attention has been paid to the rapid spectral changes that occur in the Soret region during the first 50 ps in MbCO and MbO2. In MbCO both the bleaching of the Soret peak (feature I) and the appearance of new deoxy-like absorption (feature II) occur instantaneously, whereas in MbO2 feature II is delayed with respect to feature I. A short-lived (approximately 12 ps) feature near 455 nm (feature III) was much more intense in MbO2 than in MbCO and was also identified in the transient spectrum of Mb. No evidence of subnanosecond geminate recombination was found in either MbCO or MbO2. These observations are consistent with a scheme in which MbO2 photodissociates through an excited state of Mb, whereas MbCO under the same conditions produces ground state Mb directly. The results and conclusions are compared with those of previous picosecond studies on these molecules and related hemoglobin derivatives.

Animals↗
PubMed1980

Geminate recombination of O2 and hemoglobin.

The photolysis of HbO2 and HbCO has been studied by measuring transient absorption spectra in the Soret region after excitation with picosecond pulses at 530 nm. Dissociation occurred promptly in both cases, followed (for HbO2) by geminate recombination of ca. 40% of the photodissociated O2 with a lifetime of 200 +/- 70 psec (25 degrees C). No recombination of Hb + CO was observed up to 1200 psec after photolysis. The HbO2 and HbCO photoproduct spectra were broader, weaker, and red-shifted in comparison to the spectrum of stable Hb and Gibson's fast-reacting form, Hb. For HbO2 the spectrum was initially much broader to longer wavelengths but relaxed to a constant shape within 90 psec, whereas for HbCO there was no spectral evolution. The photophysics is analyzed by considering the effect of spin constraints as well as spin--orbit coupling and orbital correlation among the various electronic states of liganded and deoxy hemoglobins. The small quantum yield of HbO2 dissociation is not primarily due to rebinding but rather to electronic relaxation to nonreactive states.

Carboxyhemoglobin↗
PubMed1980

Femtosecond relaxation of an iron porphyrin observed with polarization spectroscopy in a three-level system.

An experimental and theoretical study is presented of the nonlinear response of a dilute solution being pumped by two laser fields having different frequencies omega(1) and omega(2) that are both in resonance with electronic transitions of the solute. Experimental results were obtained for the dispersion of the third-order susceptibility chi((3))(omega(2))(2) for a dilute solution of iron(III) tetraphenylporphyrin chloride in chloroform with a fixed intense field (omega(1)) resonant with the Q(o) transition and a tunable weaker field (omega(2)) resonant with the Soret transition. The observed Lorentzian dispersion is slightly narrower than the linear absorption spectrum. The theoretical model for the third-order polarization incorporates three levels coupled to two electromagnetic fields and a population reservoir. The equations of motion for the density operator were solved in third order for the appropriate Fourier component. The resulting dispersion curve depends on both total dephasing and population decay rates, thereby demonstrating a general method for the determination of subpicosecond relaxation processes in systems to which the model is applicable. Theoretical fits to the results for iron(III) tetraphenylporphyrin chloride imply that the Soret transition is essentially homogeneously broadened with a total dephasing width of 900 +/- 200 cm(-1), and it is argued that the population decay time in this case is approximately 3.5 fsec. The broad linear absorption spectrum of the Soret band is interpreted as resulting from intramolecular perturbations between the Soret and lower energy excited states of iron(III) tetraphenylporphyrin chloride.

Journal Article↗
PubMed1978

Spectroscopic studies of oxy- and carbonmonoxyhemoglobin after pulsed optical excitation.

The photolysis of HbO2 and HbCO has been investigated with picosecond laser techniques. Transient absorption spectra were measured in the Soret and visible regions after excitation with 353- or 530-nm pulses. The photoproducts appeared within 8 psec and exhibited considerably broadened deoxyhemoglobin-like spectra, which persisted to 680 psec. The altered spectra are attributed to the production of deoxyheme conformational and spin states that might result from the intense excitation.

Carboxyhemoglobin↗