The dynamics of conformational isomerization in flexible biomolecules. I. Hole-filling spectroscopy of N-acetyl tryptophan methyl amide and N-acetyl tryptophan amide

The conformational isomerization dynamics of N-acetyl tryptophan methyl amide (NATMA) and N-acetyl tryptophan amide (NATA) have been studied using the methods of IR-UV hole-filling spectroscopy (HFS) and IR-induced population transfer spectroscopy (IR-PTS), which were developed for this purpose. Single conformations of these molecules were selectively excited in well-defined NH stretch fundamentals. This excess energy was used to drive conformational isomerization. By carrying out the infrared excitation early in a supersonic expansion, the excited molecules were recooled into their zero-point levels, partially refilling the hole created in the ground state population of one of the conformers, and creating gains in population in other conformers. These changes in population were detected using laser-induced fluorescence downstream in the expansion. In HFS, the IR wavelength is fixed and the UV laser tuned in order to determine where the population went following selective infrared excitation. In IR-PTS, the UV is fixed to monitor the population of a given conformation, and the IR is tuned to record the IR-induced changes in the population of the monitored conformer. Besides demonstrating the capability of the experiment to change the downstream conformational population distribution, the IR-PTS scans were used to extract two quantitative results: (i) The fractional populations of the conformers in the absence of the infrared, and (ii) the isomerization quantum yields for each of the six unique amide NH stretch fundamentals (three conformers each with two amide groups). The method for obtaining quantum yields is described in detail. In both NATMA and NATA, the quantum yields show modest conformational specificity, but only a hint of vibrational mode specificity. The prospects for the hole-filling technique for providing insight into energy flow in large molecules are discussed, leaving a more detailed theoretical modeling to the adjoining paper [Evans et al. J. Chem. Phys. 120, 148 (2004)].     

Laser probes of conformational isomerization in flexible molecules and complexes

Molecules with several flexible coordinates have potential energy surfaces with a large number of minima and many transition states separating them. A general experimental protocol is described that is capable of studying conformational isomerization in such circumstances, measuring the product quantum yields following conformation-specific infrared excitation, and measuring energy thresholds for isomerization of specific X --> Y reactant-product isomer pairs following excitation via stimulated emission pumping (SEP). These methods have been applied to a series of molecules of varying size and conformational complexity, including 3-indolepropionic acid (IPA), meta-ethynylstyrene, N-acetyltryptophan methyl amide (NATMA), N-acetyltryptophan amide (NATA), and melatonin. Studies of isomerization in solute-solvent complexes are also described, including a measurement of the barrier to isomerization in the IPA-H2O complex, and a unique isomerization reaction in which a single water molecule is shuttled between H-bonding sites on the trans-formanilide (TFA) molecule.     

Influence of vibrational energy flow on isomerization of flexible molecules: incorporating non-Rice-Ramsperger-Kassel-Marcus kinetics in the simulation of dipeptide isomerization

The conformational isomerization of a dipeptide, N-acetyl-tryptophan methyl amide (NATMA), is studied computationally by including important dynamical corrections to Rice-Ramsperger-Kassel-Marcus (RRKM) theory for the transition rate between pairs of isomers. The dynamical corrections arise from incomplete or sluggish vibrational energy flow in the dipeptide, a property suggested by the mode-selective chemistry that has been observed by Dian et al. [J. Chem. Phys. 120, 133 (2004)]. We compute the extent and rate of vibrational energy flow in NATMA quantum mechanically using local random matrix theory, which we then use to correct the RRKM theory rates. The latter rates are then introduced into a master equation to study the population dynamics of the dipeptide. Incomplete or slow vibrational energy flow is found to enhance the conformational selectivity of NATMA over RRKM estimates.     

Conformational preferences of jet-cooled melatonin: probing trans- and cis-amide regions of the potential energy surface

The hormone melatonin (N-acetyl-5-methoxytryptamine) is an indole derivative with a flexible peptide-like side chain attached at the C3 position. Using a combination of two-color resonant two-photon ionization (2C-R2PI), laser-induced fluorescence excitation (LIF), resonant ion-dip infrared spectroscopy (RIDIRS), fluorescence-dip infrared spectroscopy (FDIRS), and UV-UV hole-burning spectroscopy, the conformational preferences of melatonin in a molecular beam have been determined. Three major trans-amide conformers and two minor cis-amide conformers have been identified in the R2PI spectrum and characterized with RIDIRS and FDIRS. Structural assignments are made using the infrared spectra in concert with density functional theory and localized MP2 calculations. Observation of cis-amide melatonin conformers in the molecular beam, despite the large energy gap (approximately 3 kcal/mol) between trans- and cis-amides, is striking because there are at least nine lower-energy trans-amide minima that are not detected. The implications of this observation for cooling and trapping conformational population in a supersonic expansion are discussed.     

Direct measurement of the energy thresholds to conformational isomerization in tryptamine: experiment and theory

The methods of stimulated emission pumping-hole filling spectroscopy (SEP-HFS) and stimulated emission pumping population transfer spectroscopy (SEP-PTS) were applied to the conformation-specific study of conformational isomerization in tryptamine [TRA, 3-(2-aminoethyl)indole]. These experimental methods employ stimulated emission pumping to selectively excite a fraction of the population of a single conformation of TRA to well-defined ground-state vibrational levels. This produces single conformations with well-defined internal energy, tunable over a range of energies from near the zero-point level to well above the lowest barriers to conformational isomerization. When the SEP step overcomes a barrier to isomerization, a fraction of the excited population isomerizes to form that product. By carrying out SEP excitation early in a supersonic expansion, these product molecules are subsequently cooled to their zero-point vibrational levels, where they can be detected downstream with a third tunable laser that probes the ground-state population of a particular product conformer via a unique ultraviolet transition using laser-induced fluorescence. The population transfer spectra (recorded by tuning the SEP dump laser while holding the pump and probe lasers fixed) exhibit sharp onsets that directly determine the energy thresholds for conformational isomerization in a given reactant-product conformer pair. In the absence of tunneling effects, the first observed transition in a given X-Y PTS constitutes an upper bound to the energy barrier to conformational isomerization, while the last transition not observed constitutes a lower bound. The bounds for isomerizing conformer A of tryptamine to B(688-748 cm(-1)), C(1)(860-1000 cm(-1)), C(2)(1219-1316 cm(-1)), D(1219-1282 cm(-1)), E(1219-1316 cm(-1)), and F(688-748 cm(-1)) are determined. In addition, thresholds for isomerizing from B to A(<1562 cm(-1)), B to F(562-688 cm(-1)), and out of C(2) to B(<747 cm(-1)) are also determined. The A-->B and B-->A transitions are used to place bounds on the relative energies of minima B relative to A, with B lying at least 126 cm(-1) above A. The corresponding barriers have been computed using both density functional and second-order many-body perturbation theory methods in order to establish the level of theory needed to reproduce experimental results. While most of the computed barriers match experiment well, the barriers for the A-F and B-F transitions are too high by almost a factor of 2. Possible reasons for this discrepancy are discussed.     

Time-resolved IR spectroscopy of N-methylthioacetamide: trans-->cis isomerization upon n-pi and pi-pi excitation and cis-->trans photoreaction

Time-resolved infrared spectroscopy was used to study the photoisomerization of N-Methylthioacetamide (NMTAA) in D2O in both the cis-->trans and the trans-->cis direction upon selective excitation of the n-pi (S1) and pi-pi (S2) electronic transitions. While isomerization and the return to the ground state takes place on two distinct time scales (cis isomerization is 30-40%, independent of the electronic state excited, while the cis-->trans isomerization proceeds with a 60-70% quantum efficiency. These results support a mechanism by which isomerization takes place via one common intermediate state independent of electronic excitation energy and initial conformation.     

Conformational space of the Pseudosaccharin allyl ether 3-(allyloxy)-1,2-benzisothiazole 1,1-dioxide in gas phase and in rare gas matrices

In this work, the conformational space of the pseudosaccharyl ether 3-(allyloxy)-1,2-benzisothiazole 1,1-dioxide (ABID) has been studied by means of infrared spectroscopy and density functional theory (DFT) calculations. Five different low energy conformers (TSk, TC, GSk, GSk' and GC, with relative energies of 0.00, 1.97, 2.00, 3.82 and 6.02 kJ mol(-1), respectively) were found on the DFT(B3LYP)/6-311++G(3df,3pd) potential energy surface of the molecule, all of them differing in the conformation of the allyl substituent. According to the calculations, in the gaseous phase all conformers are significantly populated (TSk:TC:GSk:GSk':GC = 47%:16%:18%:12%:7%, at 350 K). In the cryogenic matrices, however, only the TSk and TC conformers exist due to isomerization from the higher energy gauche forms to the most stable trans isomers during deposition of the matrix (conformational cooling). The observed conformational cooling is in consonance with the low calculated energy barriers for the GSk --> TSk, GSk' --> TSk and GC --> TC isomerization processes. Results from annealing experiments in krypton matrix doubtlessly show that in this matrix the order of stability of the TSk and TC conformers is reversed, with the more planar TC form becoming the most stable conformer.     

Ab initio studies of conformational properties of dimethyl diphosphate dianion and its complex with magnesium

The conformational properties of the diphosphate linkage have been studied with ab initio methods using the dimethyl diphosphate dianion (1) and magnesium dimethyl diphosphate (2) as models. The ab initio energy and geometry of the conformers around the P-O bonds have been determined at the self-consistent-field (SCF) using the 6-31G* and the tzp basis sets; whereas, the 6-31G* basis set alone has been used for 2. In addition, the adiabatic connection method (ACM) of density functional theory (DFT) using the dzvp basis set has been employed for 1. The optimization of all possible staggered conformers assumed for the four P-O bonds, led to nine minima for 1. In agreement with the general anomeric effect, the sc conformation about the P-O bonds is clearly preferred over the ap one. Vibrational frequencies were calculated at the SCF level using the 6-31G* basis set and used to evaluate zero-point energies, thermal energies, and entropies for all minima of 1. The effect of zero-point energies and thermal energies is quite small. However, the effect of entropies, mainly resulting from a multiplicity contribution, changes the stability of the conformers. For each minimum of 1, up to six different arrangements of the Mg²⁺ were used to determine minima of 2. This procedure led to 21 distinct minima. The presence of the magnesium counter-ion appeared to completely change the structure and relative energy of the conformers. The preferred structures of the complex exhibit the (sc, ap) orientation around the two central P–O bonds and an arrangement in which the magnesium cation is coordinated by three phosphoryl oxygen atoms. The results of this work clearly demonstrate that interactions with the metal counter-ion can induce conformational changes in the overall 3D-shape adopted by molecules containing diphosphate linkages. The PM3 and MNDO quantum semi-empirical methods and molecular mechanics methods using the CVFF force field were tested and large differences in the minimum structures, as well as in the conformational energies between these and ab initio methods, are discussed.     

Conformational transitions of glycine induced by vibrational excitation of the O-H stretch

Vibrational energy flow and conformational transitions following excitation of the OH stretching mode of the most stable conformer of glycine are studied by classical trajectories. "On the fly" simulations with the PM3 semiempirical electronic structure method for the potential surface are used. Initial conditions are selected to correspond to the ν=1 excitation of the OH stretch. The main findings are: (1) An an equilibrium-like ratio is established between the populations of the 3 lowest-lying conformers after about 10 picoseconds. (2) There is a high probability throughout the 150 ps of the simulations for finding the molecule in geometries far from the equilibrium structures of the lowest-energy conformers. (3) Energy from the initial excited OH (ν=1) stretch flows preferentially to 5 other vibrational modes, including the bending motion of the H atom. (4) RRK theory yields conformational transition rates that deviate substantially from the classical trajectory results. Possible implication of these results for vibrational energy flow and conformational transitions in small biological molecules are discussed.     

Conformational analysis of melatonin at Hartree–Fock ab initio level

Melatonin is a hormone, which is synthesized and secreted by the pineal gland during darkness and has been implicated in the regulation of several neural and endocrine processes. Conformational analysis of melatonin was performed at Hartree–Fock ab initio level. With the help of STO-3G and 6-31G* basis sets, 192 and 128 conformers were obtained, respectively. In both cases, there were eight conformers with nearly planar 3-amido-side chain and the most stable conformation found had folded structure in accord with previously published results. The present paper compares the Hartree–Fock conformational space of melatonin with the results of active conformation models and conformational analyses published in the literature.     

Rotational isomers of small molecules in noble-gas solids: From monomers to hydrogen-bonded complexes

Molecular conformation, quantum tunneling, and hydrogen bonding play important roles in various photochemical processes. We have studied a number of small molecules possessing rotational isomerism (HONO, formic acid, acetic acid, etc.) isolated in noble-gas solid matrices. Selective vibrational excitation efficiently promotes the conformational change in the excited molecule, which allows preparation of higher-energy conformers. Stability of the higher-energy conformers is often limited by quantum tunneling of hydrogen as observed for some carboxylic acids (formic, acetic, etc.). The tunneling mechanism is supported by the strong H/D isotope effect and characteristic temperature dependence with a clear low-temperature limit. The reaction barrier height is an important factor in a tunneling process; however, other factors also play an essential role. The energy mismatch between the initial state of the higher-energy conformer and accepting state of the ground-state conformer is probably important. Hydrogen bonding can change tunneling decay rate of unstable conformers. The trans–cis formic acid dimer was prepared by vibrational excitation of the trans–trans form in neon and argon matrices. Tunneling decay of cis formic acid is substantially slower in the dimeric form compared to monomer, especially in solid neon. This stabilization effect is explained by a complexation-induced increase of reaction barrier, which is confirmed computationally. The complex between cis formic acid and water was prepared in an argon matrix and found to be stable at low-temperatures. These results show that intrinsically unstable conformational structures can be thermodynamically stabilized in asymmetrical hydrogen-bonded network. This effect occurs when the energy difference between conformers is smaller than the hydrogen bond interaction energy, which allows chemistry of unstable conformers to be studied.     

Infrared photodissociation of a water molecule from a flexible molecule-H2O complex: rates and conformational product yields following XH stretch excitation

Infrared-ultraviolet hole-burning and hole-filling spectroscopies have been used to study IR-induced dissociation of the tryptamine.H2O and tryptamine.D2O complexes. Upon complexation of a single water molecule, the seven conformational isomers of tryptamine collapse to a single structure that retains the same ethylamine side chain conformation present in the most highly populated conformer of tryptamine monomer. Infrared excitation of the tryptamine.H2O complex was carried out using a series of infrared absorptions spanning the range of 2470-3715 cm-1. The authors have determined the conformational product yield over this range and the dissociation rate near threshold, where it is slow enough to be measured by our methods. The observed threshold for dissociation occurred at 2872 cm-1 in tryptamine.H2O and at 2869 cm-1 in tryptamine.D2O, with no dissociation occurring on the time scale of the experiment (approximately 2 micros) at 2745 cm-1. The dissociation time constants varied from approximately 200 ns for the 2869 cm-1 band of tryptamine.D2O to approximately 25 ns for the 2872 cm-1 band of tryptamine.H2O. This large isotope dependence is associated with a zero-point energy effect that increases the binding energy of the deuterated complex by approximately 190 cm-1, thereby reducing the excess energy available at the same excitation energy. At all higher energies, the dissociation lifetime was shorter than the pulse duration of our lasers (8 ns). At all wavelengths, the observed products in the presence of collisions are dominated by conformers A and B of tryptamine monomer, with small contributions from the other minor conformers. In addition, right at threshold (2869 cm-1), tryptamine.D2O dissociates exclusively to conformer A in the absence of collisions with helium, while both A and B conformational products are observed in the presence of collisions with helium. Using resolution-of-identity approximation to second-order Moller-Plesset binding energies extrapolated to the complete basis set limit and harmonic vibrational frequencies and transition states calculated at the density functional limit B3LYP/6-31+G* level of theory, Rice-Ramsperger-Kassel-Marcus (RRKM) predictions for the dissociation, isomerization, and water shuttling rates as a function of energy are made. At threshold, the experimental dissociation rate is almost 10(3) faster than RRKM predictions. Reasons for this apparent non-RRKM behavior will be discussed.     

Effects of Fluorine Substitution on the Shape of Neurotransmitters: the Rotational Spectrum of 2‐(2‐Fluorophenyl)Ethanamine

The effects of fluorination on the conformational landscape of adrenergic neurotransmitters is exemplified trough the conformation analysis of 2‐(2‐F‐phenyl)ethanamine (2FPEA) carried out by microwave spectroscopy and quantum chemical calculations. Five different conformers of the nine possible stable ones for 2FPEA are observed by molecular‐beam Fourier‐transform microwave spectroscopy. Their unambiguous identification is possible by comparing the experimental rotational constants and the quadrupole coupling constants with those obtained by quantum chemical calculations carried out at the MP2/6‐311++G(d,p) level of theory. The relative abundances of the conformers in the jet are estimated from the relative intensities in the observed spectra. A qualitative agreement between experimental and theoretical energies was found, and the remaining deviations are explained by population transfer taking place during the adiabatic expansion. The energy landscape, which also takes the interconversion barriers between the conformers into consideration, is thus characterized completely by the strong interplay of quantum chemical methods and precise experimental data. Significant changes in energy and structure of the 2FPEA conformers are found compared to those obtained for the prototype molecule 2‐phenylethanamine (PEA).     

Photodissociation dynamics of hydroxybenzoic acids

Aromatic amino acids have large UV absorption cross-sections and low fluorescence quantum yields. Ultrafast internal conversion, which transforms electronic excitation energy to vibrational energy, was assumed to account for the photostability of amino acids. Recent theoretical and experimental investigations suggested that low fluorescence quantum yields of phenol (chromophore of tyrosine) are due to the dissociation from a repulsive excited state. Radicals generated from dissociation may undergo undesired reactions. It contradicts the observed photostability of amino acids. In this work, we explored the photodissociation dynamics of the tyrosine chromophores, 2-, 3- and 4-hydroxybenzoic acid in a molecular beam at 193 nm using multimass ion imaging techniques. We demonstrated that dissociation from the excited state is effectively quenched for the conformers of hydroxybenzoic acids with intramolecular hydrogen bonding. Ab initio calculations show that the excited state and the ground state potential energy surfaces change significantly for the conformers with intramolecular hydrogen bonding. It shows the importance of intramolecular hydrogen bond in the excited state dynamics and provides an alternative molecular mechanism for the photostability of aromatic amino acids upon irradiation of ultraviolet photons.     

Development of a conformational search strategy for flexible ligands: A study of the potent μ-selective opioid analgesic fentanyl

Summary An extensive conformational search of the potent opioid analgesic, fentanyl, was performed using the semiempirical quantum mechanical method AM1 and the CHARMm potential energy function. A combination of two procedures was used to search the conformational space for fentanyl, which included nested dihedral scans, geometry optimization and molecular dynamics simulation at different temperatures. In addition, the effect of a continuum solvent environment was taken into account by use of appropriate values for the dielectric constant in the CHARMm computations.The results of the conformational search allowed the determination of the probable conformation of fentanyl in polar and nonpolar solvents and of three candidate conformers for its bioactive form.     

Importance of entropy in the conformational equilibrium of phenylalanine: a matrix-isolation infrared spectroscopy and density functional theory study

The conformational behavior and infrared spectrum of l-phenylalanine were studied by matrix-isolation infrared spectroscopy and DFT [B3LYP/6-311++G(d,p)] calculations. The fourteen most stable structures were predicted to differ in energy by less than 10 kJ mol(-1), eight of them with abundances higher than 5% at the temperature of evaporation of the compound (423 K). Experimental results suggest that six conformers contribute to the spectrum of the isolated compound, whereas two conformers (IIb(3) and IIIb(3)) relax in matrix to a more stable form (IIb(2)) due to low energy barriers for conformational isomerization (conformational cooling). The two lowest-energy conformers (Ib(1), Ia) differ only in the arrangement of the amino acid group relative to the phenyl ring; they exhibit a relatively strong stabilizing intramolecular hydrogen bond of the O-H...N type and the carboxylic group in the trans configuration (O=C-O-H dihedral angle ca. 180 degrees ). Type II conformers have a weaker H-bond of the N-H...O=C type, but they bear the more favorable cis arrangement of the carboxylic group. Being considerably more flexible, type II conformers are stabilized by entropy and the relative abundances of two conformers of this type (IIb(2) and IIc(1)) are shown to significantly increase with temperature due to entropic stabilization. At 423 K, these conformers are found to be the first and third most abundant species present in the conformational equilibrium, with relative populations of ca. 15% each, whereas their populations could be expected to be only ca. 5% if entropy effects were not taken into consideration. Indeed, phenylalanine can be considered a notable example of a molecule where entropy plays an essential role in determining the relative abundance of the possible low-energy conformational states and then, the thermodynamics of the compound, even at moderate temperatures. Upon UV irradiation (lambda > 235 nm) of the matrix-isolated compound, unimolecular photodecomposition of phenylalanine is observed with production of CO(2) and phenethylamine.     

On the Influence of Higher Excited States on the ISC Quantum Yield of Octa-aL-alkyloxy-substituted Zn-Phthalocyanine Molecules Studied by Nonlinear Absorption

Abstract— Octa-aL-alkyloxy-substituted Zn-phthalocyanines are an interesting class of far red-absorbing photosensitizers. The chemical structure, the calculated steric conformation, the observed linear optical properties and an anomalous luminescence from a higher than S, excited state are reported. To study the optical properties of higher excited states and their occupation dynamics up to delay times of 15 ns we have carried out measurements of transient absorption spectra after 14 ps pulsed, resonant B-band and Q-band excitation. From these measurements the excited state singlet-singlet and triplet-triplet spectra as well as the intersystem crossing (ISC) quantum yields are obtained. The main result is an excitation wavelength-dependent ISC quantum yield that can be explained by an additional ISC channel between higher excited singlet and triplet states. The large rate of this channel is justified by the resonance between higher triplet states, observed in the triplet-triplet spectrum and the B, absorption band. Using kinetic model calculations, a lifetime of the higher excited singlet state of some picoseconds is predicted and the influence of a two-step absorption process on the population density of this higher excited singlet state is discussed.     

Conformational analysis of methylboronic acid and its acyclic esters

With the use of RHF/6-31G(d) and MP2/6-31G(d)//RHF/6-31G(d) semi-empirical (AMI) and non-empirical quantum chemical approximations conformational isomerization of methylboronic acid and its methyl, isopropyl, tert-butyl, and phenyl esters is studied. With the exception of the tert-butyl analogue, the potential energy surface of molecules in these compounds, is shown to contain three minima that correspond to planar and near-planar conformers: cis-cis, trans-trans, and cis-trans, with the latter being the main one. The minima are separated by two conformational isomerization barriers corresponding to the orthogonal arrangement of one of the OH(OR) groups.     

Conformers and photochemistry of propyl nitrites: a matrix isolation study

The infrared spectra of both constitutional isomers (n and i) of propyl nitrite have been recorded in an Ar matrix. Conformational analysis and assignments of the vibrational transitions have been carried out on the basis of quantum chemical calculations. Assignment of spectral lines to different conformers was also aided experimentally, by utilizing the different rate of photodecomposition of the conformers, as well as by employing conformational cooling using a supersonic jet as the inlet source for matrix deposition. The rate of photodecomposition is primarily determined by the steric alignment of the nitrite group, whereas jet cooling affects mainly the conformation of the alkyl tail. On the basis of these experimental observations and computational predictions two to three conformers of isopropyl nitrite and eight conformers of n-propyl nitrite were identified. After broadband ultraviolet-visible (UV-vis) photolysis of isopropyl nitrite in the matrix, HNO, acetone, HNO.acetone complex, acetaldehyde, and nitrosomethane were identified as the main products. Furthermore, in a small amount, NO and possibly the isopropoxy radical were also present in the matrix. Photolysis of n-propyl nitrite yielded HNO, propanal, and their 1:1 complex as the main products together with a small amount of NO and cis-1-nitrosopropanol.