Coded Amino Acids in Gas Phase: The Shape of Isoleucine Alberto Lesarri, Raquel Sa ´ nchez, Emilio J. Cocinero, Juan C. Lo ´ pez, and Jose ´ L. Alonso* Contribution from the Grupo de Espectroscopı ´a Molecular (GEM), Departamento de Quı ´mica Fı ´sica y Quı ´mica Inorga ´ nica, Facultad de Ciencias, UniVersidad de Valladolid, 47005 Valladolid, Spain Received April 29, 2005; E-mail: jlalonso@qf.uva.es Abstract: The solid R-amino acid isoleucine has been vaporized by laser ablation and expanded in a supersonic jet, where the molecular conformations of the isolated molecule were probed using Fourier transform microwave spectroscopy. Two conformers of neutral isoleucine have been detected in gas phase, the most stable being stabilized by an intramolecular hydrogen bond N-H‚‚‚OdC and a cis-COOH arrangement. The higher energy form is stabilized by an intramolecular hydrogen bond N‚‚‚H-O. The sec-butyl side chain of the amino acid adopts the same configuration in the two observed conformers, with a staggered configuration at C similar to that observed in valine and a trans arrangement of CR and Cδ. Ab initio calculations at MP2/6-311++G(d,p) level reproduce satisfactorily the experimental results. Introduction The dynamic role of R-amino acids (NH 2 -CH(R)-COOH) as building blocks of proteins relies on their high torsional flexibility, which results in a large number of low energy conformational minima. The preferred conformations are de- termined by a delicate balance of different covalent and noncovalent interactions within the molecule and with its surroundings, especially hydrogen bonding. In particular, amino acids in crystals or solutions are stabilized as charge-separated zwitterions 1 (NH 3 + -CH(R)-COO - ) by a network of inter- molecular hydrogen bond interactions. As a consequence, the intramolecular interactions and the intrinsic conformational preferences of these systems cannot be determined in condensed phases and are only revealed when the molecules are isolated in gas phase, where the amino acids exhibit an unsolvated neutral form (NH 2 -CH(R)-COOH). This form represents the best approximation to the electronic environment of an amino acid residue in a polypeptide chain or protein. A supersonic jet expansion with an inert carrier gas is the preferred experimental approach to isolate the different conformers in their separated potential wells. 2 The strong collisional regime at the beginning of the adiabatic expansion produces a strong cooling of the rotational and vibrational states, and the individual conformers are usually frozen into the ground vibrational state of each individual well. In this way, the conformer distribution before the expansion may be preserved provided that interconversion barriers between conformers are sufficiently high. As the expansion evolves, the number of molecular collisions practi- cally disappears, and thus the spectroscopic properties of the different species can be probed in a local environment of virtual isolation. Different experimental methodologies have been used for the study in gas phase of amino acids and other small bioactive molecules. 3 Electronic spectroscopy of amino acids is limited to favorable cases that present aromatic chromophores. 4,5 The electronic spectrum of the different conformers can be analyzed with the aid of double resonance techniques (UV-UV, 6 IR- UV 7 ) and ab initio theoretical predictions, but do not provide direct structural information since in most cases rotational resolution is not attainable. Rotational spectroscopy is the only technique that, thanks to its inherently superior resolution, can distinguish unambiguously between different isomers, conform- ers, or isotopomers and provide accurate structural information directly comparable to the in vacuo theoretical predictions. However, amino acids are difficult to vaporize since they are solids with high melting points and thermally unstable. For these reasons, most neutral amino acids have escaped for a long time to gas-phase spectroscopic investigation, and even the intrinsic conformational landscape of the genetically encoded amino acids is poorly known. The analysis of the rotational spectrum of natural amino acids, started in the late 1970s by Suenram and Godfrey on glycine 8,9 and later by Godfrey on alanine 10 and (1) (a) Albrecht, G.; Corey, R. B. J. Am. Chem. Soc. 1939, 61, 1087. (b) Marsh, R. E. Acta Crystallogr. 1958, 11, 654. (c) Levy, H. A.; Corey, R. B. J. Am. Chem. Soc. 1941, 63, 2095. (d) Donohue, J. J. Am. Chem. Soc. 1950, 72, 949. (2) (a) Levy, D. H. Annu. ReV. Phys. Chem. 1980, 31, 197. (b) Jet Spectroscopy and Molecular Dynamics; Hollas, J. M., Phillips, D., Eds.; Blackie: London, 1995. (3) (a) Robertson, E. G.; Simons, J. P. Phys. Chem. Chem. Phys. 2001, 3, 1. (b) Weinkauf, R.; Schermann, J.-P.; de Vries, M. S.; Kleinermans, K. Eur. Phys. J. D 2002, 20, 309. (c) Zwier, S. J. Phys. Chem. A 2001, 105, 8827. (4) Snoek, L. C.; Robertson, E. G.; Kroemer, R. T.; Simons, J. P. Chem. Phys. Lett. 2000, 321, 49. (5) Bakker, J. M.; Aleese, L. M.; Meijer, G.; von Helden, G. Phys. ReV. Lett. 2003, 91, 203003. (6) Lipert, R. J.; Colson, S. D. Chem. Phys. Lett. 1998, 161, 303. (7) (a) Page, R. H.; Shen, Y. R.; Lee, Y. T. J. Chem. Phys. 1988, 88, 4621. (b) Page, R. H.; Shen, Y. R.; Lee, Y. T. J. Chem. Phys. 1988, 88, 5362. (8) (a) Suenram, R. D.; Lovas, F. J. J. Mol. Spectrosc. 1978, 72, 372. (b) Suenram, R. D.; Lovas, F. J. J. Am. Chem. Soc. 1980, 102, 7180. (c) Lovas, F. J.; Kawashima, Y.; Grabow, J.-U.; Suenram, R. D.; Fraser, G. T.; Hirota, E. Astrophys. J. 1995, 455, L201. Published on Web 08/23/2005 12952 9 J. AM. CHEM. SOC. 2005, 127, 12952-12956 10.1021/ja0528073 CCC: $30.25 © 2005 American Chemical Society