3D equilibrium crystal shapes of Pb: the significance of step-resolved images

Scanning tunnelling microscopy has proven to be an extremely useful technique for imaging small crystallites equilibrated at elevated temperature. As an example, we review recent work on three-dimensional Pb crystallites of 1–2 μm diameter, supported on Ru(0 0 1). Large (1 1 1) facets and, depending on temperature, small (1 1 2) facets were observed in the top section of the crystallites. The temperature dependent facet anisotropy was analyzed to yield the absolute step free energies of Pb(1 1 1) vicinal steps. The vicinal region close to the (1 1 1) facet was studied in detail to determine the shape exponent, the step–step interaction energy and the constant of the dipole interaction potential. Boundary conditions of the evaluated vicinal region have been specified for proving the universality of shapes, characterized by the exponent of 3/2, which is clear evidence for the 1/x² step interaction potential. The role of the activation barrier for facet growth or shrinkage is discussed in the context of attaining 3D equilibrium of crystallites. A comparative study of crystallites with defect-free and dislocated facets shows significant differences, providing direct evidence of the activation barrier. Reliable step–step interaction energies were obtained for dislocated crystallites. Extrapolating the temperature dependent total step interaction energy to 0 K yields for the first time values of the structure dependent dipole–dipole step interaction energies of A- and B-steps.     

Selective IR-induced isomerization of 1,2-dichloropropane isolated in xenon matrix

The conformations of monomeric 1,2-dichloropropane isolated in low temperature xenon matrix were characterized experimentally using FTIR spectroscopy. The interpretation of the experimental spectra was aided by ab initio MP2/6-311++G(3df,3pd) calculations. Relative energies of three structures, differing by the orientation of the Cl−C−C−Cl dihedral angle, fall within 0–6 kJ mol⁻¹ range. The spectral signatures of all three conformers were identified experimentally. Infrared irradiation of the matrix isolated compound resulted in selective conversion of the most stable Trans conformer into the higher energy Gauche+ rotamer, while the Gauche− conformer remained unaffected. This result was interpreted in terms of calculated energy barriers and dipole moments.     

Microwave and vibrational spectra, ab initio calculations, conformational stabilities and assignments of the fundamentals of the Cs conformer of n-butylgermane

The FT-microwave spectrum of n-butylgermane, CH₃CH₂CH₂CH₂GeH₃ has been investigated from 4000 to 18,000 MHz and the microwave spectra have been observed for all of the five naturally occurring germanium isotopologues for the anti-anti (aa) conformer. The dipole moment for the ⁷⁴Ge containing species has been measured, giving a total dipole moment of 0.881 (26) D. In addition, the vibrational spectrum of n-butylgermane is described. Modestly complete assignments are made for the aa conformer. The relative stabilities of the five conformers are calculated, and the anti-anti (aa) conformer is found to be the most stable in all calculations done. This conclusion is confirmed by the infrared and Raman spectrum of the annealed crystal. The dipole moments of all conformers are calculated to be approximately equal and less than 1 D, ranging from approximately 0.8 to 0.9 D.     

Conformational identification of tryptamine embedded in superfluid helium droplets using electronic polarization spectroscopy

We report electronic polarization spectroscopy of tryptamine embedded in superfluid helium droplets. In a dc electric field, dependence of laser induced fluorescence from tryptamine on the polarization direction of the excitation laser is measured. Among the three observed major conformers A, D, and E, conformers D and E display preference for perpendicular excitation relative to the orientation field, while conformer A is insensitive to the polarization direction of the excitation laser. We attribute the behavior of conformer A to the fact that the angle between the permanent dipole and the transition dipole is close to the magic angle. Using a linear variation method, we can reproduce the polarization preference of the three conformers and determine the angle between the transition dipole and the permanent dipole. Since the side chain exerts small effect on the direction of the transition dipole in the frame of the indole chromophore, all three conformers have a common transition dipole more or less in the indole plane at an angle of approximately 60 degrees relative to the long axis of the chromophore. The orientation of the side chain, on the other hand, determines the size and direction of the permanent dipole, thereby affecting the angle between the permanent dipole and the transition dipole. For conformer D in the droplet, our results agree with the Anti(ph) structure, rather than the Anti(py) structure. Our work demonstrates that polarization spectroscopy is effective in conformational identification for molecules that contain a known chromophore. Although coupling of the electronic transition with the helium matrix is not negligible, it does not affect the direction of the transition dipole.     

Gas Phase Conformations of Tetrapeptide Glycine-Phenylalanine-Glycine-Glycine

Systematic search of the potential energy surface of tetrapeptide glycine-phenylalanine-glycine-glycine (GFGG) in gas phase is conducted by a combination of PM3, HF and BHandHLYP methods. The conformational search method is described in detail. The rela-tive electronic energies, zero point vibrational energies, dipole moments, rotational constants, vertical ionization energies and the temperature dependent conformational distributions for a number of important conformers are obtained. The structural characteristics of these conformers are analyzed and it is found that the entropic effect is a dominating factor in determining the relative stabilities of the conformers. The measurements of dipole moments and some characteristic IR mode are shown to be effective approaches to verify the theoreti-cal prediction. The structures of the low energy GFGG conformers are also analyzed in their connection with the secondary structures of proteins. Similarity between the local structures of low energy GFGG conformers and the α-helix is discussed and many β- and γ-turn local structures in GFGG conformers are found.     

Electron energy loss spectroscopy of liquid glycerol

The inelastic scattering of low energy electrons from liquid glycerol has been studied. For the first time, electron energy loss spectra of liquids are sufficiently well resolved to permit the identification of vibrations corresponding to individual bonds, namely the C–H and O–H stretching vibrations in glycerol. The angular distribution of the specular peak is very broad, indicating the absence of long-range order at the surface of the liquid. The measurement of the loss signals as a function of the primary electron energy suggests a hybrid mechanism of excitation. The excitation mechanism for the O–H vibration has a stronger impact character as compared to the C–H vibration. A negative ion resonance of glycerol is found at a primary energy of 8 eV. The signal intensities measured as a function of the specular angle of the electron beam appear to be influenced by the angular dependence of the dipole and impact scattering cross-section and a possible preferred orientation of the C–H and O–H groups at the surface of the liquid.     

Raman and infrared spectra, conformational stability, ab initio calculations and vibrational assignments for 2-chloroethyl silane

The infrared (3500–30 cm⁻¹) spectra of gaseous and solid and the Raman (3500–10 cm⁻¹) spectra of liquid with quantitative depolarization ratios and solid 2-chloroethyl silane, ClCH₂CH₂SiH₃, have been recorded. Similar data have been recorded for the Si–d₃ isotopomer. These data indicate that two conformers, trans and gauche, are present in the fluid states but only one conformer, trans, is present in the solid. The mid-infrared spectra of the sample dissolved in liquified xenon as a function of temperature (−55 to −100°C) has been recorded. The enthalpy difference between the conformers has been determined to be 181±12 cm⁻¹ (2.17±0.14 kJ/mol) with the trans rotamer the more stable form. From the isolated Si–H frequencies from the Si–d₂ isotopomer the r_o Si–H distances of 1.484 and 1.483 Å for the trans and 1.481 for the gauche conformers have been obtained. Ab initio calculations have been carried out with several different basis sets up to MP2/6-311+G** from which structural parameters and conformational stabilities have been determined. With all the basis sets the trans form is predicted to be the more stable conformer which is consistent with the experimental results. These results are compared to the corresponding quantities for the carbon analogue.     

Charge redistribution on electronic excitation. Dipole moments of cis- and trans-3-aminophenol in their S(0) and S(1) electronic states

Rotationally resolved electronic spectroscopy in the gas phase, in the absence and presence of an applied electric field, has been used to distinguish the two conformers of 3-aminophenol (3AP) on the basis of differences in their electric dipole moments. cis-3AP has micro = 2.3 D, and trans-3AP has micro = 0.7 D, in their ground electronic states. The two observed values are approximately equal to those expected on the basis of bond dipole additivity rules. However, these rules fail to predict the large change in both the magnitude and the orientation of micro when the two conformers of 3AP absorb light. cis-3AP has micro = 3.3 D, and trans-3AP has micro = 1.7 D, in their excited S(1) electronic states; the angles of orientation of micro with respect to the a inertial axis change by 13 degrees and 38 degrees, respectively. This effect is attributed to (1)L(b)/(1)L(a) state mixing in the S(1) state.     

Novel inhibitor discovery through virtual screening against multiple protein conformations generated via ligand-directed modeling: a maternal embryonic leucine zipper kinase example

Kinase targets have been demonstrated to undergo major conformational reorganization upon ligand binding. Such protein conformational plasticity remains a significant challenge in structure-based virtual screening methodology and may be approximated by screening against an ensemble of diverse protein conformations. Maternal embryonic leucine zipper kinase (MELK), a member of serine-threonine kinase family, has been recently found to be involved in the tumerogenic state of glioblastoma, breast, ovarian, and colon cancers. We therefore modeled several conformers of MELK utilizing the available chemogenomic and crystallographic data of homologous kinases. We carried out docking pose prediction and virtual screening enrichment studies with these conformers. The performances of the ensembles were evaluated by their ability to reproduce known inhibitor bioactive conformations and to efficiently recover known active compounds early in the virtual screen when seeded with decoy sets. A few of the individual MELK conformers performed satisfactorily in reproducing the native protein-ligand pharmacophoric interactions up to 50% of the cases. By selecting an ensemble of a few representative conformational states, most of the known inhibitor binding poses could be rationalized. For example, a four conformer ensemble is able to recover 95% of the studied actives, especially with imperfect scoring function(s). The virtual screening enrichment varied considerably among different MELK conformers. Enrichment appears to improve by selection of a proper protein conformation. For example, several holo and unliganded active conformations are better to accommodate diverse chemotypes than ATP-bound conformer. These results prove that using an ensemble of diverse conformations could give a better performance. Applying this approach, we were able to screen a commercially available library of half a million compounds against three conformers to discover three novel inhibitors of MELK, one from each template. Among the three compounds validated via experimental enzyme inhibition assays, one is relatively potent (15; K(d) = 0.37 μM), one moderately active (12; K(d) = 3.2 μM), and one weak but very selective (9; K(d) = 18 μM). These novel hits may be utilized to assist in the development of small molecule therapeutic agents useful in diseases caused by deregulated MELK, and perhaps more importantly, the approach demonstrates the advantages of choosing an appropriate ensemble of a few conformers in pursuing compound potency, selectivity, and novel chemotypes over using single target conformation for structure-based drug design in general.     

Searching and optimizing structure ensembles for complex flexible sugars

NMR restrictions are suitable to specify the geometry of a molecule when a single well-defined global free energy minimum exists that is significantly lower than other local minima. Carbohydrates are quite flexible, and therefore, NMR observables do not always correlate with a single conformer but instead with an ensemble of low free energy conformers that can be accessed by thermal fluctuations. In this communication, we describe a novel procedure to identify and weight the contribution to the ensemble of local minima conformers based on comparison to residual dipolar couplings (RDCs) or other NMR observables, such as scalar couplings. A genetic algorithm is implemented to globally minimize the R factor comparing calculated RDCs to experiment. This is done by optimizing the weights of different conformers derived from the exhaustive local minima conformational search program, fast sugar structure prediction software (FSPS). We apply this framework to six human milk sugars, LND-1, LNF-1, LNF-2, LNF-3, LNnT, and LNT, and are able to determine corresponding population weights for the ensemble of conformers. Interestingly, our results indicate that in all cases the RDCs can be well represented by only a few most important conformers. This confirms that several, but not all of the glycosidic linkages in histo-blood group "epitopes" are quite rigid.     

A semi-empirical and ab initio combined approach for the full conformational searches of gaseous lysine and lysine–H2O complex

A full structural search of the canonical, zwitterionic, protonated and deprotonated lysine conformers in gas phase is presented. A total of 17,496 canonical, 972 zwitterionic, 11,664 protonated and 1458 trial deprotonated structures were generated by allowing for all combinations of internal single-bond rotamers. All the trial structures were initially optimized at the AM1 level, and the resulting structures were determined at the B3LYP/6-311G^* level. A total of 927 canonical, 730 protonated and 193 deprotonated conformers were found, but there were no stable zwitterionic structures in the gas phase. The most stable conformers of the canonical, protonated and deprotonated lysine were further optimized at the B3LYP/6-311++G^** level. The energies of the most stable structures were determined at the MP2/6-311G(2df,p) level and the vibrational frequencies were calculated at the B3LYP/6-311++G^** level. The rotational constants, dipole moments, zero-point vibrational energies, harmonic frequencies, vertical ionization energies, enthalpies, Gibbs free energies and conformational distributions of gaseous lysine were presented. Numerous new structures are found and the lowest-energy lysine conformer is more stable than the existing one by 1.1 kcal/mol. Hydrogen bonds are classified and may cause significant red-shifts to the associated vibrational frequencies. The calculated proton affinity/dissociation energy and gas-phase basicity/acidity are in good agreement with the experiments. Calculations are also presented for the canonical lysine–H₂O and zwitterionic lysine–H₂O clusters. Interaction between lysine and H₂O significantly affects the relative conformational stabilities. Only one water molecule is sufficient to produce the stable zwitterionic structures in gas phase. The lowest-energy structure is found to be zwitterions when applying the conductor-like polarized continuum solvent model (CPCM) to the lysine–H₂O complexes.     

Probing dynamic protein ensembles with atomic proximity measures

The emerging role of internal dynamics in protein fold and function requires new avenues of structure analysis. We analyzed the dynamically restrained conformational ensemble of ubiquitin generated from residual dipolar coupling data, in terms of protruding and buried atoms as well as interatomic distances, using four proximity-based algorithms, CX, DPX, PRIDE and PRIDE-NMR (http://hydra.icgeb.trieste.it/protein/). We found that Ubiquitin, this relatively rigid molecule has a highly diverse dynamic ensemble. The environment of protruding atoms is highly variable across conformers, on the other hand, only a part of buried atoms tends to fluctuate. The variability of the ensemble cautions against the use of single conformers when explaining functional phenomena. We also give a detailed evaluation of PRIDE-NMR on a wide dataset and discuss its usage in the light of the features of available NMR distance restraint sets in public databases.     

Nonlinear dielectric effect (NDE) and molecular orbital study of the conformational equilibrium in 1,4-dimethoxybenzene in benzene solution

The conformational equilibrium in 1,4-dimethoxybenzene (1,4-DMB) in benzene solutions has been studied. On the basis of experimental values of the nonlinear dielectric effect (NDE) parameter, electric permittivity and density, determined in this work, and applying the general statistical theory of NDE, the contributions of the syn-anti and syn-syn conformers and the electric dipole moment of the polar syn-syn conformer were calculated. The molecular orbital method (PM3) has also been applied for calculation of the dipole moments and energies of particular conformers. The results of the NDE study and PM3 calculations are consistent and they reveal the existence of two conformers (syn-anti and syn-syn) of comparable energy values, but different values of dipole moments, and the predominance of the polar form (syn-syn) of the mole fraction in benzene. Moreover, the energies of intermolecular interactions have been determined from the concentration dependence of linear and nonlinear polarisability.     

A microwave and quantum chemical study of the conformational properties and intramolecular hydrogen bonding of 1-fluorocyclopropanecarboxylic acid

The structural and conformational properties of 1-fluorocyclopropanecarboxylic acid have been explored by microwave spectroscopy and a series of ab initio (MP2/6-311++G(d,p) level), density functional theory (B3LYP/aug-cc-pVTZ level), and G3 quantum chemical calculations. Four "stable" conformers, denoted conformers I-IV, were found in the quantum chemical calculations, three of which (conformers I -III) were predicted to be low-energy forms. Conformer I was in all the quantum chemical calculations predicted to have the lowest energy, conformer III to have the second lowest energy, and conformer II to have the third lowest energy. Conformers II and III were calculated to have relatively large dipole moments, while conformer I was predicted to have a small dipole moment. The microwave spectrum was investigated in the 18-62 GHz spectral range. The microwave spectra of conformers II and III were assigned. Conformer I was not assigned presumably because its dipole moment is comparatively small. Conformer II is stabilized by an intramolecular hydrogen bond formed between the fluorine atom and the hydrogen atom of the carboxylic acid group. Conformer III has a synperiplanar orientation for the F-C-C=O and H-O-C=O chains of atoms. Its dipole moment is: mua = 3.4(10), mub = 10.1(13), and muc = 0.0 (assumed) and mu(tot) = 10.6(14) x 10(-30) C m [3.2(4) D]. Several vibrationally excited states of the lowest torsional mode of each of II and III were also assigned. The hydrogen-bonded conformer II was found to be 2.7(2) kJ/mol less stable than III by relative intensity measurements. Absolute intensity measurements were used to show that the unassigned conformer I is the most abundant form present at a concentration of roughly 65% at room temperature. Conformer I was estimated to be ca. 5.0 kJ/mol more stable than the hydrogen-bonded rotamer (conformer II) and ca. 2.3 kJ/mol more stable than conformer III. The best agreement with the theoretical calculations is found in the MP2 calculations, which predict conformer I to be 5.1 kJ/mol more stable than III and 1.7 kJ/mol more stable than II.     

Far infrared spectra, conformational equilibria, vibrational assignments, ab initio calculations and structural parameters for 2-bromoethanol

The far infrared spectrum from 370 to 50 cm⁻¹ of gaseous 2-bromoethanol, BrCH₂CH₂OH, was recorded at a resolution of 0.10 cm⁻¹. The fundamental O–H torsion of the more stable gauche (Gg′) conformer, where the capital G refers to internal rotation around the C–C bond and the lower case g to the internal rotation around the C–O bond, was observed as a series of Q-branch transitions beginning at 340 cm⁻¹. The corresponding O–H torsional modes were observed for two of the other high energy conformers, Tg (285 cm⁻¹) and Tt (234 cm⁻¹). The heavy atom asymmetric torsion (rotation around C–C bond) for the Gg′ conformer has been observed at 140 cm⁻¹. Variable temperature (−63 to −100°C) studies of the infrared spectra (4000–400 cm⁻¹) of the sample dissolved in liquid xenon have been recorded. From these data the enthalpy differences have been determined to be 411±40 cm⁻¹ (4.92±0.48 kJ/mol) for the Gg′/Tt and 315±40 cm⁻¹ (3.76±0.48 kJ/mol) for the Gg′/Tg, with the Gg′ conformer the most stable form. Additionally, the infrared spectrum of the gas, and Raman spectrum of the liquid phase are reported. The structural parameters, conformational stabilities, barriers to internal rotation and fundamental frequencies have been obtained from ab initio calculations utilizing different basis sets at the restricted Hartree–Fock or with full electron correlation by the perturbation method to second order. The theoretical results are compared to the experimental results when appropriate. Combining the ab initio calculations with the microwave rotational constants, r₀ adjusted parameters have been obtained for the three 2-haloethanols (F, Cl and Br) for the Gg′ conformers.     

The electronic structure and conformation of dimethylaminobenzonitrile

Ab initio molecular orbital (MO) computations on the closed shell singlet ground state ofN, N-dimethylaminobenzonitrile (DMABN) are reported. Fully optimized structures of several conformers of DMABN were calculated at the HF/6-31G level of theory. Our results indicate that for each of these conformations the minimum energy structure has a trigonal (sp² hybridized] amino nitrogen. The most stable DMABN conformer was found to be planar with its methyl groups eclipsed. The Koopmans ionization potentials and dipole moments of the various ground state conformers are compared. The implications for dynamical models of twisted-intramolecular charge transfer (TICT) are discussed. Moreover, the use of qualitative MO theory arguments provides an interpretation of the computational results in a simple orbital interaction framework.A. Mellon Foundation Fellow (1989–1992).Operated for the U.S. Department of Energy by Battelle Memorial Institute under Contract DE-AC06-76RLO 1830.     

Infrared and Raman spectra, conformational stability, ab initio calculations, and vibrational assignments for methylaminothiophosphoryl difluoride

Infrared spectra (4000–50 cm⁻¹) of the vapor, amorphous and crystalline solids and Raman spectra (3600–10 cm⁻¹) of the liquid with qualitative depolarization data as well as the amorphous and crystalline solids of methylaminothiophosphoryl difluoride, CH₃N(H)P(=S)F₂, and three deuterated species, CD₃N(H)P(=S)F₂, CH₃N(D)P(=S)F₂, and CD₃N(D)P(=S)F₂, have been recorded. The spectra indicate that in the vapor, liquid and amorphous solid a small amount of a second conformer is present, whereas only one conformer remains in the low temperature crystalline phase. The near-infrared spectra of the vapor confirms the existence of two conformers in the gas phase. Asymmetric top contour simulation of the vapor shows that the trans conformer is the predominant vapor phase conformer. From a temperature study of the Raman spectrum of the liquid the enthalpy difference between the trans and near-cis conformers was determined to be 368±15 cm⁻¹ (4.41±0.2 kJ/mol), with the trans conformer being thermodynamically preferred. Ab Initio calculations with structure optimization using the 6-31G(d) and 6-311+G(d,p) basis sets at the restricted Hartree–Fock (RHF) and/or with full electron correlation by the perturbation method to second order (MP2) support the occurrence of near-trans (5° from trans) and near-cis (20° from cis) conformers. From the RHF/6-31G(d) calculation the near-trans conformer is predicted to be the more stable form by 451 cm⁻¹ (5.35 kJ/mol) and from the MP2/6-311+G(d,p) calculation by 387 cm⁻¹ (4.63 kJ/mol). All of the normal modes of the near-trans rotamer have been assigned based on infrared band contours, depolarization values and group frequencies and the assignment is supported by the normal coordinate calculation utilizing harmonic force constants from the MP2/6-31G(d) ab initio calculations.     

Conformational and structural studies of 1-fluorosilacyclobutane from temperature dependent FT-IR spectra of krypton solutions and ab initio calculations

The infrared spectra (3500 to 40 cm⁻¹) of gaseous and solid and the Raman spectra (3500 to 30 cm⁻¹) of liquid and solid 1-fluorosilacyclobutane, c-C₃H₆SiFH, have been obtained. Both the axial and equatorial conformers with respect to the fluorine atom have been identified in the fluid phases. Variable temperature (−105 to −150 °C) studies of the infrared spectra of the sample dissolved in liquid krypton have been carried out. From these data, the enthalpy difference has been determined to be 282 ± 27 cm⁻¹ (3.37 ± 0.32 kJ/mol), with the equatorial conformer the more stable form and the only conformer remaining in the annealed solid. At ambient temperature there is approximately 21 ± 2% of the axial conformer present in the vapor phase. From isolated Si–H stretching frequencies the Si–H (r₀) distances are calculated to be 1.484 and 1.485 Å for the equatorial and axial conformers, respectively. Structural parameters have been predicted from MP2/6-311 + G(d,p) ab initio calculations and the adjusted r₀ parameters for both conformers were obtained from a combination of the ab initio predicted values and the six previously reported microwave rotational constants. Along with the Si–H bond distance, the Si–C, and C–C distances of 1.865(5), and 1.571(5) Å, respectively, for the equatorial conformer are significantly different from the values for these parameters previously reported from an election diffraction study. Both the SiC and CC distances and the SiF distance are longer by 0.002 and 0.004 Å, respectively, for the axial conformer. Structural parameters have also been obtained for silacyclobutane, c-C₃H₆SiH₂ and ethylsilylfluoride, CH₃CH₂SiH₂F, from combined ab initio predicted values and previously reported rotational constants. Several of these newly determined parameters are significantly different from those previously reported for both molecules. Complete equilibrium geometries, conformational stabilities, harmonic force fields, infrared intensities, Raman activities, and depolarization ratios have been determined for both rotamers by ab initio calculations employing the 6-31G(d) basis set at the level of Moller–Plesset (MP) to second order. A complete vibrational assignment supported by normal coordinate calculations is proposed for the equatorial conformer, and several of the fundamentals of the axial conformer have also been identified. The results are discussed and compared to corresponding quantities for some similar molecules.     

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 Spectroscopy of 2-Furoic Acid and Its Dimer: Conformational Distribution and Double Proton Tunneling

Structural and tunneling properties of the 2-furoic acid (FA) monomer and dimer were investigated using rotational spectroscopy and DFT calculations. CREST, a conformational ensemble space exploration tool, was used to identify all possible low-energy conformations of the FA monomer and dimer, followed by the DFT geometry optimization and harmonic frequency calculations. Broadband rotational spectra in the 2–6 and 8–12 GHz regions were recorded in a supersonic jet expansion. The monomeric FA was found to exist dominantly as three different conformers: I , II , and III in a jet, with I and II taking on the cis-COOH configuration while III having the trans-COOH configuration. For the FA dimer, only the I – II conformer was observed experimentally, whereas the symmetric I – I and II – II conformers were not observed because of their zero dipole moments. The analysis of the splittings in the rotational transitions of I – II allowed one to extract the tunneling splitting to be 1056.0(12) MHz. The barrier height was determined to be ∼442 cm⁻¹ using the scaled potential energy scans at several different levels of theory.