Mid-Infrared Study of Deoxyadenosine at High Pressures: Evidence of Phase Transitions

Abstract

Room temperature mid-infrared experiments between 500 and 1800 cm^?1 have been performed on crystalline deoxyadenosine as a function of pressure up to about 10 GPa. Discontinuities observed near 2 and 4 GPa indicate that two separate phase transitions occur at these pressures. Changes in the spectra suggest that both transitions involve a rearrangement of the pucker of the deoxyribose moiety. The wavenumbers of the vibrational modes shift to higher values with applied pressure. Our results for deoxyadenosine are compared to similar measurements on adenosine. Assignments for the observed modes are made on the basis of work published in the literature.     

Raman and infrared studies of nucleosides at high pressures: I. Adenosine.

Room temperature Raman and infrared (IR) spectra of crystalline adenosine at pressures between 1 atm and 10 GPa are reported. Vibrational modes were identified through assignments in the literature. Many modes were found to increase in frequency with pressure; however, some irregularities were observed. Discontinuities were observed in numerous Raman and IR modes near 2.5 GPa, indicating a phase transition. The modes associated with the glycosidic bond shift significantly down in frequency near this pressure, suggesting a weakening of the associated bond. The IR modes associated with hydrogen-stretching motions were found to decrease in frequency with pressure.     

Raman and Infrared Studies of Nucleosides at High Pressures: I. Adenosine

Abstract

Room temperature Raman and infrared (IR) spectra of crystalline adenosine at pressures between 1 atm and 10 GPa are reported. Vibrational modes were identified through assignments in the literature. Many modes were found to increase in frequency with pressure; however, some irregularities were observed. Discontinuities were observed in numerous Raman and IR modes near 2.5 GPa, indicating a phase transition. The modes associated with the glycosidic bond shift significantly down in frequency near this pressure, suggesting a weakening of the associated bond. The IR modes associated with hydrogen-stretching motions were found to decrease in frequency with pressure.     

Mid-infrared study of deoxycytidine at high pressures: evidence of a phase transition

Room temperature mid-infrared experiments between 600 and 1600 cm(-1) have been performed on crystalline deoxycytidine as a function of pressure up to about 10 GPa. The wavenumbers of most vibrational modes shift to higher values with increasing pressures except for a mode near 840 cm(-1). Assignments for the observed modes are made on the basis of work published in the literature. Several anomalies are noted near 4.7 GPa, suggesting a phase transition. Our results for deoxycytidine are compared to similar measurements on cytidine.     

The effects of high pressure upon ligated and deoxyhemoglobins and myoglobin. An optical spectroscopic study

The effects of high pressure (0.1-3.4 gigapascal (GPa)) on the ferrous heme active sites of human adult hemoglobin, sperm whale myoglobin, and Glycera dibranchiata hemoglobin (Fraction II) were probed using resonance Raman and absorption spectroscopies. High-to-low spin transitions of the heme iron occur for hemoglobin, myoglobin, and Glycera hemoglobin at 0.35, 0.75, and 0.50 GPa, respectively, for the deoxy species. These interspecies differences result from variations in the composition of the hemepockets and/or their rigidity to pressure-induced volume changes. Heme active sites initially bound to CO or O2 exhibit distinctive behavior at high pressures. For all proteins studied, O2 apparently dissociates from the heme at only moderately high pressure, while CO remains bound to the heme moiety even at extreme pressures. The Raman spectra demonstrate the differences in the ligated and deoxy species at 3.4 GPa in the high frequency region. Discrete changes (i.e. iron spin-state transitions and dissociation of O2) occur that are commensurate with the collapse of the distal pocket, while continuous shifts in the absorption and Raman spectra are observed at pressures above those required for pocket collapse.     

Raman and infrared studies of nucleosides at high pressures: II. Cytidine

Raman and mid-infrared (MIR) spectra have been recorded for crystalline cytidine at pressures up to 10 GPa at room temperature. Broadening and positive wavenumber shifts are observed for most of the Raman and MIR peaks with increasing pressure. However, some of the MIR peaks associated with hydrogen-stretching modes display a negative wavenumber shift as a result of charge transfer effects. Evidence of a phase transition near 4 GPa is presented and attributed to a change in the conformation of the five membered sugar ring.     

1H-NMR parameters of common amino acid residues measured in aqueous solutions of the linear tetrapeptides Gly-Gly-X-Ala at pressures between 0.1 and 200 MPa

For the interpretation of chemical shift changes induced by pressure in proteins, a comparison with random-coil data is important. For providing such a data basis, the pressure dependence of the 1H-NMR chemical shifts of the amino acids X in the random-coil model peptides Gly-Gly-X-Ala were studied for the 20 common amino acids at two pH values (pH 5.0 and 5.4) at 305 K, in the pressure range from 0.1 to 200 MPa. The largest shift changes deltadelta with pressure p can be observed for the backbone amide protons. The average linear pressure coefficient delta(deltap) is 0.38 ppm GPa(-1), with a root mean square deviation of 0.2 ppm GPa(-1). In contrast to the downfield shift typical for amide protons, the H(alpha)-resonances typically shift upfield, with a pressure coefficient of -0.025 ppm GPa(-1) and a root mean square deviation of 0.05 ppm GPa(-1). The side chain resonances are only weakly influenced by pressure, on average they are shifted by 0.014 ppm GPa(-1)) with a root mean square deviation of 0.14 ppm GPa(-1). The exceptions are the side chain amide protons of asparagine and glutamine. Here, values of 0.214 (Asn H(delta21)), 0.417 (Asn H(delta22)), 0.260 (Gln H(varepsilon21)) and 0.395 (Gln H(varepsilon22)) ppm GPa(-1) can be observed. In both cases, the pressure dependent shift is larger for the pro-E proton than for the pro-Z proton. Within the limits of error the equilibrium constant for the trans- and cis-conformers at the proline peptide bond is independent of pressure in the pressure range studied.     

Excited-State Lifetimes of Far-Infrared Collective Modes in Proteins

Vibrational excitations of low frequency collective modes are essential for functionally important conformational transitions in proteins. Here we report the first direct measurement on the lifetime of vibrational excitations of the collective modes at 87 pm (115 cm^-1) in bacteriorhodopsin, a transmembrane protein. The data show that these modes have extremely long lifetime of vibrational excitations, over 500 picoseconds, accommodating 1500vibrations. We suggest that there is a connection between this relativelyslow anharmonic relaxation rate of approximately 10 g sec^-1 and thesimilar observed rate of conformational transitions in proteins, which require require multi-level vibrational excitations and energy exchanges with othervibrational modes and collisional motions of solvent molecules.     

Energy landscape of a native protein: Jumping-among-minima model

We have investigated energy landscape of human lysozyme in its native state by using principal component analysis and a model, jumping-among-minima (JAM) model. These analyses are applied to 1 nsec molecular dynamics trajectory of the protein in water. An assumption embodied in the JAM model allows us to divide protein motions into intra-substate and inter-substate motions. By examining intra-substate motions, it is shown that energy surfaces of individual conformational substates are nearly harmonic and mutually similar. As a result of principal component analysis and JAM model analysis, protein motions are shown to consist of three types of collective modes, multiply hierarchical modes, singly hierarchical modes, and harmonic modes. Multiply hierarchical modes, the number of which accounts only for 0.5% of all modes, dominate contributions to total mean-square atomic fluctuation. Inter-substate motions are observed only in a small-dimensional subspace spanned by the axes of multiplyhierarchical and singly hierarchical modes. Inter-substate motions have two notable time components: faster component seen within 200 psec and slower component. The former involves transitions among the conformational substates of the low-level hierarchy, whereas the latter involves transitions of the higher level substates observed along the first four multiply hierarchical modes. We also discuss dependence of the subspace, which contains conformational substates, on time duration of simulation. Proteins 33:496–517, 1998. © 1998 Wiley-Liss, Inc.     

The effect of slow allosteric transitions in a coupled biochemical oscillator model.

The effect of slowed allosteric transitions in a coupled biochemical oscillator model showing complex dynamic behavior is investigated. When the allosteric transitions are sufficiently fast one can obtain a low-dimensional asymptotic approximation for the dynamics of the species that evolve on a slow time-scale. Such low-dimensional models are common in studies of biological control systems and little attention has, so far, been given to the dynamic effect of the large number of species usually eliminated from more biochemically detailed models. Here we investigate the dynamic effect of explicit inclusion of allosteric transitions having finite time-scales of equilibration. It is found that slowed allosteric transitions suppress complex dynamic modes such a bursting, quasi-periodicity and chaos. The effect arises as the enzyme of consideration becomes trapped in an active state where it is unable to respond to changes in effector concentration on the time-scale necessary to support the modes of complex dynamics. Slow allosteric transitions may be favourable in biological systems in which complex oscillations are not desirable but which, at the same time, may benefit from the presence of positive feedbacks. Our findings suggest that slow allosteric transitions and finite internal rates in general may contribute significantly to the dynamics of biological control mechanisms.     

Stability diagram and unfolding of a modified cytochrome c: what happens in the transformation regime?

We determined the stability diagram of a modified cytochrome c protein in a glycerol water mixture by measuring the first and the second moment of the fluorescence from the chromophore as a function of temperature and pressure. Temperature and pressure were varied between 273 and 363 K and 0.0001 and 1 GPa, respectively. The shift of the fluorescence maximum showed a characteristic sigmoid-like pattern from which information on the microscopic processes during unfolding is obtained: as the transformation regime is entered, the fluorescence shows a significant blue shift. The conclusion is that water molecules get into contact with the chromophore. They lead to strong electrostatic contributions in the solvent shift, which counteract the red shifting dispersion interactions. Assuming that there are just two relevant states that determine the stability diagram, the complete set of thermodynamic parameters can be determined from the data. However, under certain pressure-temperature conditions the fluorescence pattern is more complicated, pointing toward reentrant transitions and, possibly, to consecutive steps in the unfolding process.     

The decisive role of the water structure in changes of conformation of nucleic acids.

This review summarizes data on the structure and properties of water under normal conditions, at high salt concentration and under high pressure. We correlate the observed conformational transitions in nucleic acids with changes in water structure and activity, and suggest a mechanism of conformational transitions of nucleic acid involving these changes. We conclude that the Z-DNA form is induced only at low water activity caused by high salt concentrations and/or high pressure.     

A Raman study of the hydration of wet-spun films of Li-hyaluronate

Raman spectroscopy between 620 and 1700 cm(-1) has been performed on wet-spun films of Li-hyaluronate as a function of relative humidity between 54 and 93% at room temperature. The observed vibrational modes show no significant dependence on water content, suggesting that the molecule does not undergo any conformational transitions in this humidity range.     

On the propagation of a wave front in viscoelastic arteries

In formulating a mathematical model of the arterial system, the one-dimensional flow approximation yields realistic pressure and flow pulses in the proximal as well as in the distal regions of a simulated arterial conduit, provided that the viscoelastic damping induced by the vessel wall is properly taken into account. Models which are based on a purely elastic formulation of the arterial wall properties are known to produce shocklike transitions in the propagating pulses which are not observed in man under physiological conditions. The viscoelastic damping characteristics are such that they are expected to reduce the tendency of shock formation in the model. In order to analyze this phenomenon, the propagation of first and second-order pressure waves is calculated with the aid of a wave front expansion, and criteria for the formation of shocks are derived. The application of the results to the human arterial system show that shock waves are not to be expected under normal conditions, while in case of a pathologically increased pressure rise at the root of the aorta, shocklike transitions may develop in the periphery. In particular, it is shown that second-order waves never lead to shock formation in finite time for the class of initial conditions and mechanical wave guides which are of interest in the mammalian circulation.     

S-state dependent Fourier transform infrared difference spectra for the photosystem II oxygen evolving complex

Vibrational spectroscopy provides a means to investigate molecular interactions within the active site of an enzyme. We have applied difference FTIR spectroscopy coupled with a flash turnover protocol of photosystem II (PSII) to study the oxygen evolving complex (OEC). Our data show two overlapping oscillatory patterns as the sample is flashed through the four-step S-state cycle that produces O(2) from two H(2)O molecules. The first oscillation pattern of the spectra shows a four-flash period four oscillation and reveals a number of new vibrational modes for each S-state transition, indicative of unique structural changes involved in the formation of each S-state. Importantly, the first and second flash difference spectra are reproduced in the 1800-1200 cm(-)(1) spectral region by the fifth and sixth flash difference spectra, respectively. The second oscillation pattern observed is a four-flash, period-two oscillation associated with changes primarily to the amide I and II modes and reports on changes in sign of these modes that alternate 0:0:1:1 during S-state advance. This four-flash, period-two oscillation undergoes sign inversion that alternates during the S(1)-to-S(2) and S(3)-to-S(0) transitions. Underlying this four-flash period two is a small-scale change in protein secondary structure in the PSII complex that is directly related to S-state advance. These oscillation patterns and their relationships with other PSII phenomena are discussed, and future work can initiate more detailed vibrational FTIR studies for the S-state transitions providing spectral assignments and further structural and mechanistic insight into the photosynthetic water oxidation reaction.     

Changing of the dynamics modes in populations with age and sex structure

Using a mathematical model of the number dynamics in a population with age and sex structure, the relationships were studied between the number of newborns and that of mature females and males. In addition, we studied transitions between different modes of population dynamics which were dependent on changes in not only the population parameters, such as reproduction intensity, survival, and self-regulation, but also in mating pair formation. Population dynamics was found to manifest multistability; i.e., the alternative dynamics modes were observed at the same values of the population parameters but under different initial conditions.     

Plasticity of tumor cell migration: acquisition of new properties or return to the past?

During tumor development cancer cells pass through several stages when cell morphology and migration abilities change remarkably. These stages are named epithelial-mesenchymal and mesenchymal-amoeboid transitions. The molecular mechanisms underlying cell motility are changing during these transitions. As result of transitions the cells acquire new characteristics and modes of motility. Cell migration becomes more independent from the environmental conditions, and thus cell dissemination becomes more aggressive, which leads to formation of distant metastases. In this review we discuss the characteristics of each of the transitions, cell morphology, and the specificity of cellular structures responsible for different modes of cell motility as well as molecular mechanisms regulating each transition.     

First-principles study of the structural transformation, electronic structure, and optical properties of crystalline 2,6-diamino-3,5-dinitropyrazine-1-oxide under high pressure

Periodic first-principles calculations have been performed to study the effect of high pressure on the geometric, electronic, and absorption properties of 2,6-diamino-3,5-dinitropyrazine-1-oxide (LLM-105) under hydrostatic pressures of 0–50 GPa. Obvious irregular changes in lattice constants, unit-cell angles, bond lengths, bond angles, and band gaps showed that crystalline LLM-105 undergoes four structural transformations at 8, 17, 25, and 42 GPa, respectively. The intramolecular H-bonds were strong at pressures of 0–41 GPa but weakened in the range 42–50 GPa. The lengths of the intermolecular H-bonds (<1.47 Å) indicated that these H-bonds have covalent character and tend to induce the formation of a new twelve-membered ring. Analysis of the DOS showed that the interactions between electrons, especially the valence electrons, strengthen under the influence of pressure. The p states play a very important role in chemical reactions of LLM-105. The absorption spectrum of LLM-105 displayed more bands—as well as stronger bands—in the fundamental absorption region when the pressure was high rather than low. A new absorption peak due to O–H stretching appeared at 18.3 eV above 40 GPa, indicating that covalent O–H bonds and a new twelve-membered ring are present in LLM-105.     

Vibrational analysis and spectra of orotic acid

The IR and Raman spectra of polycrystalline anhydrous orotic acid and its N1, N3, and O12 trideuterated isotopomer are recorded in the 4000-40 cm(-1) spectral interval as part of a series of vibrational analyses of nucleosides, nucleotides, and related compounds carried out in our laboratory. The frequencies of the fundamental transitions and the potential energy distributions of the 39 normal modes of orotic acid and its isotopomer are calculated by an ab initio density functional theory Becke3P86/6-311G** treatment. Assignments of the vibrational modes are proposed that consider the results of these calculations and the observed spectra. The results of the ab initio treatment are related to crystallographic and spectral data, and they are compared with previous assignments for similar molecules.     

Low-frequency vibrational modes and infrared absorbance of red, blue and green opsin

Vibrational excitations of low-frequency collective modes are essential for functionally important conformational transitions in proteins. We carried out an analysis of the low-frequency modes in the G protein coupled receptors (GPCR) family of cone opsins based on both normal-mode analysis and molecular dynamics (MD) simulations. Power spectra obtained by MD can be compared directly with normal modes. In agreement with existing experimental evidence related to transmembrane proteins, cone opsins have functionally important transitions that correspond to approximately 950 modes and are found below 80 cm⁻¹. This is in contrast to bacteriorhodopsin and rhodopsin, where the important low-frequency transition modes are below 50 cm⁻¹. We find that the density of states (DOS) profile of blue opsin in a solvent (e.g. water) has increased populations in the very lowest frequency modes (<15 cm⁻¹); this is indicative of the increased thermostability of blue opsin. From our work we found that, although light absorption behaves differently in blue, green and red opsins, their low-frequency vibrational motions are similar. The similarities and differences in the domain motions of blue, red and green opsins are discussed for several representative modes. In addition, the influence of the presence of a solvent is reported and compared with vacuum spectra. We thus demonstrate that terahertz spectroscopy of low-frequency modes might be relevant for identifying those vibrational degrees of freedom that correlate to known conformational changes in opsins. An erratum to this article can be found at