Carbonyl Stretch Research Articles

Origins of the Sensitivity of Molecular Vibrations to Electric Fields: Carbonyl and Nitrosyl Stretches in Model Compounds and Proteins

The vibrational Stark effect (VSE) is the effect of an electric field on the vibrational spectrum. Typically, the largest change induced by an electric field is the linear frequency shift whose sensitivity is known as theStark tuning rate Δμ; the magnitude and direction of Δμ are measured by VSE spectroscopy. This can be used to calibrate the sensitivity of vibrational frequencies to local electrostatic fields in proteins. To be most useful, it is critical to obtain a sensitive probe that has a large Δμ, and understanding the origins of Δμ is required. The Stark tuning rate arises from two distinct origins: mechanical anharmonicity (Δμ a n h ) and the change of the bond force constant because of the electronic polarizability (Δμ b o n d ), with Δμ = Δμ a n h + Δμ b o n d . These two origins were quantitatively investigated for carbonyl and nitrosyl stretches in model compounds and proteins by IR and VSE spectroscopy. The contribution of Δμ a n h was found to dominate Δμ for carbonyl stretches in acetone, methyl vinyl ketone, and 1-methyl-2-pyrrolidinone despite their different degrees of electron delocalization. In contrast, the presence of d-π * back-bonding when CO is bound to the heme causes Δμ b o n d to be more significant, giving rise to large variations in the Fe-CO and C-O stretch frequencies in heme protein variants such as myoglobin mutants, with a strong negative correlation between v F e - C O and v C - O . The back-bonding also increases Δμ a n h because of the enhanced effective charges on the CO oscillator. By comparison, the small variation in Fe-NO stretch frequency and the absence of a strong negative correlation between Fe-NO and N-O stretch frequencies for NO bound to the heme iron in myoglobin variants suggest that Δμ F e - N O is much smaller than Δμ F e C O , possibly because of less back-bonding in heme-NO complexes. Nevertheless, Δμ N - O has approximately the same magnitude as Δμ C - O when bound to heme, suggesting that the anharmonic effect on VSE is larger for the N-O stretch than that for C-O stretch.

Origins of the Sensitivity of Molecular Vibrations to Electric Fields: Carbonyl and Nitrosyl Stretches in Model Compounds and Proteins

The vibrational Stark effect (VSE) is the effect of an electric field on the vibrational spectrum. Typically, the largest change induced by an electric field is the linear frequency shift whose sensitivity is known as the Stark tuning rate Δμ; the magnitude and direction of Δμ are measured by VSE spectroscopy. This can be used to calibrate the sensitivity of vibrational frequencies to local electrostatic fields in proteins. To be most useful, it is critical to obtain a sensitive probe that has a large Δμ, and understanding the origins of Δμ is required. The Stark tuning rate arises from two distinct origins: mechanical anharmonicity (Δμanh) and the change of the bond force constant because of the electronic polarizability (Δμbond), with Δμ = Δμanh + Δμbond. These two origins were quantitatively investigated for carbonyl and nitrosyl stretches in model compounds and proteins by IR and VSE spectroscopy. The contribution of Δμanh was found to dominate Δμ for carbonyl stretches in acetone, methyl vinyl ketone, and 1-methyl-2-pyrrolidinone despite their different degrees of electron delocalization. In contrast, the presence of d−π* back-bonding when CO is bound to the heme causes Δμbond to be more significant, giving rise to large variations in the Fe−CO and C−O stretch frequencies in heme protein variants such as myoglobin mutants, with a strong negative correlation between ν̄Fe-CO and ν̄C-O. The back-bonding also increases Δμanh because of the enhanced effective charges on the CO oscillator. By comparison, the small variation in Fe−NO stretch frequency and the absence of a strong negative correlation between Fe−NO and N−O stretch frequencies for NO bound to the heme iron in myoglobin variants suggest that ΔμFe-NO is much smaller than ΔμFe-CO, possibly because of less back-bonding in heme-NO complexes. Nevertheless, ΔμN-O has approximately the same magnitude as ΔμC-O when bound to heme, suggesting that the anharmonic effect on VSE is larger for the N−O stretch than that for C−O stretch.

Time-Resolved Resonance Raman Spectroscopic Studies on the Radical Anions of Methyl-1,4-benzoquinone and 2,6-Dimethyl-1,4-benzoquinone

Time-resolved resonance Raman spectroscopy (TR3) and density functional calculations have been used to study the effect of asymmetric substitution of methyl group on the structure and vibrational spectra of the radical anion of benzoquinone. The asymmetrically substituted benzoquinones that have been studied are methyl-1,4-benzoquinone (MBQ) and 2,6-dimethyl-1,4-benzoquinone (2,6 DMBQ). Specific vibrational mode assignments have been made to all the vibrational frequencies recorded in the experiment. From the structural changes upon reduction to the radical anion, it has been observed that the unpaired electron in the antibonding $\pi *$ orbital is more localized on the C=O bond proximal to the methyl group which results in increase in proximal carbonyl bond length compared to the distal bond. The coupling between the C=O bonds has been analyzed on the basis of the (DFT results) shift in the vibrational frequency upon isotopic substitution of carbonyl carbon and oxygen atoms. Comparison of the $(^1^8O)$ isotopic shift and shift in their C=O stretching frequencies between MBQ and 2,6 DMBQ suggests in the case of 2,6 DMBQ that coupling of the C=O modes with other internal coordinates is considerable for the symmetric C=O stretch compared to asymmetric C=O stretch. Further studies based on site-specific isotopic substitution of the carbonyl carbon and oxygen atoms suggest that symmetric C=O stretch is predominantly from the proximal carbonyl bond and the extent of contribution from the proximal carbonyl bond increases in case of 2,6 DMBQ. The asymmetric C=O stretch has slightly more contribution from the distal carbonyl stretch than the proximal.

Coupling and orientation between anharmonic vibrations characterized with two-dimensional infrared vibrational echo spectroscopy

Two-dimensional vibrational spectroscopy has been used to characterize transient molecular structure by measuring the couplings and projection angles between two strongly coupled anharmonic vibrations. Two-dimensional Fourier-transform infrared spectra of the coupled carbonyl stretches of Rh(CO)2(C5H7O2) in hexane have been obtained from femtosecond vibrational echo signals detected with spectral interferometry. The eight resonances in the two-dimensional spectrum can be interpreted as two diagonal peaks and two cross peaks, each split into a pair. The splitting between the peak pairs is directly related to the diagonal and off-diagonal anharmonicity of the symmetric and asymmetric carbonyl stretches. The ratio of the amplitude of the cross peaks for two different polarization geometries determines the projection angle between the coupled transition dipoles. The experimental characterization of the vibrational eigenstates allows the local carbonyl structure to be modeled as bilinearly coupled cubic anharmonic oscillators. The interaction between the carbonyl stretches arises from the mutual bonding with the rhodium metal center. This two-dimensional infrared experiment characterizes the structure with a time window of roughly 20 ps, suggesting a general method for capturing transient molecular structure in solution.

Ion–molecule interactions in solutions of lithium perchlorate in propylene carbonate+diethyl carbonate mixtures: an IR and molecular orbital study

FTIR spectra have been recorded and analyzed for solutions of lithium perchlorate in propylene carbonate (PC), diethyl carbonate (DEC), and PC+DEC mixtures. It has been shown that the carbonyl stretch bands for PC and DEC are very sensitive to the interaction between Li + and the solvent molecules. They split with addition of LiClO 4, indicating a strong interaction of Li + with PC and DEC through the oxygen group of PC and both oxygen and ether oxygen atoms of DEC. In conjunction with molecular orbital calculation, the optimized geometries of solvation are given. In addition, solvent separated ion pairs and contact ion pairs were observed in LiClO 4/DEC solutions, and no preferential solvation of Li + in LiClO 4/PC+DEC solutions were detected.

Raman investigation of single oxidized carbon nanotubes

AbstractThe oxidation process of single‐walled carbon nanotubes via nitric acid treatment was followed by IR‐, UV‐Vis‐NIR, and single bundle Raman spectroscopy. The introduction of functional, oxygen‐containing groups is revealed by an additional absorption band at 1725 cm−1, characteristic of carbonyl stretch vibrations. No significant shift of the optical absorption bands could be detected after oxidation. The combination of atomic force microscopy and confocal scanning resonance‐enhanced Raman microscopy was used to investigate thin bundles and, eventually, individual nanotubes in detail. These experiments enabled determination of the dependence of the Raman intensity of the G‐line (around 1590 cm−1) on the bundle height for both non‐oxidized and oxidized tubes. The Raman cross‐section of the oxidized tubes was found to be reduced by a factor of ˜4, compared to the pristine tubes. This observation is ascribed to all tubes within a bundle that are oxidized to the same degree.

Correlation Effects in the Two-Dimensional Vibrational Spectroscopy of Coupled Vibrations

Correlated energy shifts in the spectral broadening of coupled vibrational transitions are shown to have clear signatures in infrared two-dimensional dispersed vibrational echo (DVE) spectroscopy. A model that includes correlation effects through a correlation coefficient (ρ) for a bivariate distribution is used to describe two-dimensional experiments on the coupled carbonyl stretches of Rh(CO)2(C5H7O2) (or RDC) in chloroform. Signatures of correlated (0 < ρ < +1) and anticorrelated (−1 < ρ < 0) broadening in DVE experiments manifest themselves in the depth of modulation of the signal and the magnitude of the echo peak shift. For the case of RDC, the broadening is highly correlated (ρ = 0.9) and can be explained in terms of the solvent-induced modulation of the Rh electron density.

Mechanism of unfolding of a model helical peptide.

Synthetic model helical peptides, Acetyl-W(EAAAR)(5)A-amide with (13)C=O specifically labeled alanine segments in repeats n = 1,2 or 4,5 were studied in aqueous D(2)O solution as a function of temperature using Fourier transform infrared spectroscopy and two-dimensional correlation analysis. The (13)C==O provided a probe which was sensitive to the carbonyl stretch in the peptide bonds of the alanine residues at the amino terminal end in one peptide as compared to the probe in the carboxy terminal end of the other peptide during thermal perturbation. The relative stability of each terminal end was examined; the more stable terminal was determined to be the amino terminal end. Also studied were the glutamate and arginine side-chain modes involved in the salt bridging interaction. Two-dimensional correlation analysis enabled enhanced resolution in the spectral region of 1520--1700 cm(-1), and thus, the order in which these vibrational modes were perturbed as a function of increasing temperature were established.

Solvent effects on vibrational modes: ab-initio calculations, scaling and solvent functions with applications to the carbonyl stretch of dialkyl ketones

We report calculations on harmonic vibrational frequencies for the carbonyl stretch of a series of ketones of general formula R-CO-Me in various solvents. The frequencies are calculated at the B3LYP/6-31G(d) level by using recent versions of the Polarizable Continuum Model (PCM). A comparison with experimental data is carried out with the help of different strategies for the scaling of computed values in solution. It is found that a solvent-specific scaling of the standard PCM frequencies is able to reproduce the fine details in the experimental solvent shifts. A parallel comparison of experimental, non-scaled and scaled frequencies with respect to solvent functions of various nature permits to identify the cases in which specific solute–solvent interactions play a role. For such cases more refined versions of PCM, using a quantum mechanical description of the solute accompanied by a small set of solvent molecules, may be used, if there is the need of refining the standard description.

Acetic Acid Diffusion in Polyisobutylene: Probing Small Molecule Structures

In this study, time-resolved Fourier transform infrared, attenuated total reflectance (FTIR-ATR) spectroscopy was used to characterize the diffusion of acetic acid in polyisobutylene (PIB) at different vapor activities in order to understand complex diffusion mechanisms and probe molecular structures. The relationship between the carbonyl (C=O) stretch at 1715 cm -1 , which represents a cyclic dimer, and an unknown structure at 1726 cm -1 was investigated during the diffusion process to determine the structure of the unknown species. A local equilibrium association model suggested that the unknown species was a linear dimer in equilibrium with the cyclic dimer. A mathematical model for diffusion with isomerization, assuming local equilibrium, predicted that each species would have the same effective diffusion coefficient, but the individual diffusion coefficients could not be separated from the expression for the effective diffusion coefficient. Numerical simulations of the two coupled continuity equations were performed to show that the individual diffusion coefficients can be separated and that they are equal to each other for the acetic acid/PIB system. The values of the individual diffusion coefficients are consistent with the structural hypothesis from the local equilibrium assumption.

Theoretical and spectroscopic characterization of 1,4-bis-ethoxymethylene-(2,3)-butadione: a molecule with an extraordinary low frequency “CO” stretching mode

During a recent investigation of the photophysical properties of Ru(II) complexes containing the ligand 4,4′-bipyridine we had occasion to prepare an intermediate, 1,4-bisethoxymethylene-2,3-butadione, abbreviated as EMB. We were surprised to find a strong band at ca. 1590 cm −1 (IR spectra) which had all the characteristics of a carbonyl stretch but saw no comparable bands in the region where one would normally expect the CO resonance. This frequency is the lowest among all known organic carbonyl compounds. We performed theoretical calculations on isolated EMB molecule by using Gaussian 94 package (RHF and DFT methods with the 6-311G and 6-311+G(d,p) basis sets). Within the DFT methods the functionals SVWN and B3LYP have been tested. Calculated IR, Raman, 1 H NMR and 13 C NMR spectra appeared to be in good agreement with experimental data. We also calculated potential energy distribution (PED) for presented in the paper vibrational bands.

Rhodopsin Activation Affects the Environment of Specific Neighboring Phospholipids: An FTIR Spectroscopic Study

Rhodopsin is a member of a superfamily of G-protein-coupled receptors that transduce signals across membranes. We used Fourier-transform infrared (FTIR) difference spectroscopy to study the interaction between rhodopsin and lipid bilayer upon receptor activation. A difference band at 1744 cm −1 (+)/1727 cm −1 (−) was identified in the FTIR-difference spectrum of rhodopsin mutant D83N/E122Q in which spectral difference bands arising from the carbonyl stretching frequencies of protonated carboxylic acid groups were removed by mutation. As the band was abolished by detergent delipidation, we suggested that it arose from carbonyl groups of phospholipid fatty acid esters. Rhodopsin and the D83N/E122Q mutant were reconstituted into various 13C-labeled 1-palmitoyl-2-oleoyl- sn-glycero-3-phosphocholine vesicles and probed. The 1744-cm −1 (+)/1727 cm −1 (−) band could be unequivocally assigned to a change in the lipid ester carbonyl stretch upon receptor activation, with roughly equal contribution from both lipid esters. The band intensity scaled with the amount of rhodopsin but not with the amount of lipid, excluding the possibility that it was due to the bulk lipid phase. We also excluded the possibility that the lipid band represents a change in the number of boundary lipids or a general alteration in the boundary lipid environment upon formation of metarhodopsin II. Instead, the data suggest that the lipid band represents the change of a specific lipid-receptor interaction that is coupled to protein conformational changes.

Functional groups and elemental analyses of cuticular morphotypes of Cordaites principalis (Germar) Geinitz, Carboniferous Maritimes Basin, Canada

Well-preserved cuticles were isolated from Cordaites principalis (Germar) Geinitz leaf compressions, i.e., foliage from extinct gymnosperm trees Coniferophyta: Order Cordaitales. The specimens were collected from the Sydney, Stellarton and Bay St. George subbasins of the once extensive Carboniferous Maritimes Basin of Atlantic Canada. Fourier transformation of infrared spectra (FTIR) and elemental analyses indicate that the ca. 300–306-million-year-old fossil cuticles share many of the functional groups observed in modern cuticles. The similarities of the functional groups in each of the three cuticular morphotypes studied support the inclusion into a single cordaite-leaf taxon, i.e., C. principalis (Germar), confirming previous morphological investigations. Vitrinite reflectance measurements on coal seams in close proximity to the fossil-bearing sediments reveal that the Bay St. George sample site has the lowest thermal maturity, whereas the sites in Sydney and Stellarton are more mature. IR absorption and elemental analyses of the cordaite compressions corroborate this trend, which suggests that the coalified mesophyll in the leaves follows a maturation path similar to that of vitrinite. Comparison of functional groups of the cordaite cuticles with those from certain pteridosperms previously studied from the Sydney Subbasin shows that in the cordaite cuticles highly conjugated C–O (1632 cm −1) bands dominate over carbonyl stretch that characterizes the pteridosperm cuticles. The differences demonstrate the potential of chemotaxonomy as a valuable tool to assist distinguishing between Carboniferous plant–fossil groups.

Probing solvation dynamics with femtosecond vibrational spectroscopy

We explain why solvent reorganization can induce both red- and blue-shifting of vibrational transitions of a particular probe molecule upon excitation to the S1 electronic state. We observe with femtosecond vibrational spectroscopy, after hydrogen-bond cleavage dynamics, an additional blue shift of the carbonyl stretch of coumarin 102 of 7 cm-1 when dissolved in chloroform.

Enzyme-induced strain/distortion in the ground-state ES complex in beta-lactamase catalysis revealed by FTIR.

Class A beta-lactamases hydrolyze penicillins and other beta-lactams via an acyl-enzyme catalytic mechanism. Ser70 is the active site nucleophile. By constructing the S70A mutant, which is unable to form the acyl-enzyme intermediate, it was possible to make stable ES complexes with various substrates. The stability of such Michaelis complexes permitted acquisition of their infrared spectra. Comparison of the beta-lactam carbonyl stretch frequency (nu(CO)) in the free and enzyme-bound substrate revealed an average decrease of 13 cm(-)(1), indicating substantial strain/distortion of the lactam carbonyl when bound in the ES complex. Interestingly, regardless of the frequency of the C=O stretch in the free substrate, when complexed to Bacillus licheniformis beta-lactamase, the frequency was always 1755 +/- 2 cm(-)(1). This suggests the active site environment induces a similar conformation of the beta-lactam in all substrates when bound to the enzyme. Using deuterium substitution, it was shown that the "oxyanion hole", which involves hydrogen bonding to two backbone amides, is the major source of the enzyme-induced strain/distortion. The very weak catalytic activity of the S70A beta-lactamase suggests enzyme-facilitated hydrolysis due to substrate distortion on binding to the enzyme. Thus the binding of the substrate in the active site induces substantial strain and distortion that contribute significantly to the overall rate enhancement in beta-lactamase catalysis.

Do Parallel β-Helix Proteins Have a Unique Fourier Transform Infrared Spectrum?

Several polypeptides have been found to adopt an unusual domain structure known as the parallel β-helix. These domains are characterized by parallel β-strands, three of which form a single parallel β-helix coil, and lead to long, extended β-sheets. We have used ATR-FTIR (attenuated total reflectance-fourier transform infrared spectroscopy) to analyze the secondary structure of representative examples of this class of protein. Because the three-dimensional structures of parallel β-helix proteins are unique, we initiated this study to determine if there was a corresponding unique FTIR signal associated with the parallel β-helix conformation. Analysis of the amide I region, emanating from the carbonyl stretch vibration, reveals a strong absorbance band at 1638 cm −1 in each of the parallel β-helix proteins. This band is assigned to the parallel β-sheet structure. However, components at this frequency are also commonly observed for β-sheets in many classes of globular proteins. Thus we conclude that there is no unique infrared signature for parallel β-helix structure. Additional contributions in the 1638 cm −1 region, and at lower frequencies, were ascribed to hydrogen bonding between the coils in the loop/turn regions and amide side-chain interactions, respectively. A 13-residue peptide that forms fibrils and has been proposed to form β-helical structure was also examined, and its FTIR spectrum was compared to that of the parallel β-helix proteins.

Do interstrand hydrogen bonds contribute to β-hairpin peptide stability in solution? IR analysis of peptide folding in water

The amide I carbonyl stretch in the IR spectrum, together with 1H NMR Hα chemical shifts, have been used to investigate the folding of a 16-residue β-hairpin peptide in water: while Hα shifts are consistent with a significant population of the folded state (ca. 40%), we see no features in the IR spectrum in the amide I region to suggest a significant contribution from interstrand hydrogen bonds, although at high peptide concentration (10 mM) the appearance of a new band at 1616 cm−1 is consistent with the onset of irreversible peptide aggregation.

Fourier transform infrared spectra of a late intermediate of the bacteriorhodopsin photocycle suggest transient protonation of Asp-212.

We measured time-resolved difference spectra, in the visible and the infrared, for the Glu-194 and Glu-204 mutants of bacteriorhodopsin and detected an anomalous O state, labeled O', in addition to the authentic O intermediate, before recovery of the initial state in the photocycle. The O' intermediate exhibits prominent bands at 1712 cm(-1) (positive) and 1387 cm(-1) (negative). These bands arise with the same time constant as the deprotonation of Asp-85. Both bands are shifted to lower frequency upon labeling of the protein with [4-(13)C]aspartic acid. The former band, but not the latter, is shifted in D2O. These shifts identify the two bands as the carboxyl stretch of a protonated aspartic acid and the symmetric carbonyl stretch of an unprotonated aspartate, respectively, and suggest that in O' an initially anionic aspartate enters into protonation equilibrium with Asp-85. Elimination of the few other candidates, on various grounds, identifies Asp-212 as the unknown residue. It is possible, therefore, that in the last step of the photocycle of the mutants studied the proton released from Asp-85 is conducted to the extracellular surface via Asp-212. An earlier report of a weak band at 1712 cm(-1) late in the wild-type photocycle [Zscherp and Heberle (1997) J. Phys. Chem. B 101, 10542-10547] suggests that Asp-212 might play this role in the wild-type protein also.

Anion photoelectron spectroscopy of PdCO− and PdCN: Reactivity of Pd−

The ground electronic states of PdCO−/PdCO and PdCN−/PdCN are investigated using anion photoelectron spectroscopy (PES). The electron affinity of PdCO− is determined to be 0.604±0.010 eV. The anion PES of PdCO− exhibits a single, vibrationally resolved electronic band assigned to the Σ+1+e−←2Σ+ anion to neutral transition. The neutral carbonyl stretch is resolved and measured to be 2140±60 cm−1. There is an additional, partially resolved 350±40 cm−1 vibrational progression that is attributed to Pd–CO relative motion. The PES of PdCN− exhibits a single peak at 2.543±0.007 eV, which is the electron affinity of PdCN. This vertical electronic band is tentatively assigned to the Σ+2+e−←1Σ+ transition. The highest occupied molecular orbital of both the anion and neutral is described as a nonbonding spd hybrid orbital on Pd that does not affect the ionic bond of the neutral Pd+⋅CN− core. The various reactions of Pd and Pd− with methanol and acetonitrile along with other reactants that may be present in the ion source are considered.

Comparison of the Structures and Vibrational Modes of Carboxybiotin and N-Carboxy-2-imidazolidone

The spontaneous decarboxylation of N-carboxy-2-imidazolidone (a model for carboxybiotin) and N(1‘)-carboxybiotin can be followed at high pH by Raman and FTIR spectroscopies. The major bands associated with vibrations of the carboxylate group have been assigned on the basis of quantum mechanical calculations of N-carboxy-2-imidazolidone and N(1‘)-carboxy-2-methylbiotin. The carboxylate modes are the asymmetric stretch, coupled to the ureido carbonyl stretch, near 1710 cm-1, the symmetric stretch near 1340 cm-1, and the −CO2- scissoring motion near 830 cm-1. In the case of carboxybiotin, the last two modes are strongly coupled with biotin ring modes. All three carboxylate modes disappear as spontaneous decarboxylation occurs, to be replaced by features attributable to the noncarboxylated ring structures.The HF/6-31G* optimized structure of 2-methylbiotin revealed that the ureido ring portion is essentially planar, in accord with a number of X-ray crystallographic structures of biotin compounds. However, calculations at this level and at the B3LYP/6-31+G(d) level (using density functional theory) predict that the ureido ring in biotin puckers upon carboxylation. Comparison of the structures of carboxybiotin and carboxyimidazolidone, derived at the HF/6-31G* level, indicates that lengths of the ring-nitrogen-to-carboxylate bonds are equal and that the torsional angles about this linkage are very similar. This strong structural similarity provides a rationale for the observation that, at high pH, the spontaneous rates of decarboxylation of these two molecules are very similar.