Spectroscopy and dissociation dynamics of cold, biomolecular ions

Monia Guidi
EPFL, 2010
Thèse EPFL

Résumé

The function of a peptide is intimately connected to its structure, which in solution is the result of the balance between non-covalent interactions within the molecule and the molecule-solvent interactions. Theory has the power to predict the properties of a wide variety of molecules, but the accuracy of the theoretical predictions must be verified by experiments. A way to test, and hopefully improve, theory is to study proteins or peptides in the gas-phase. The combination of mass spectrometry and laser spectroscopy is used to investigate properties of biological ions in the gas-phase. Closed-shell biomolecular ions produced by an electrospray source are cooled down in an RF ion trap to less than 10 K and interrogated spectroscopically. Electronic and conformer-specific vibrational spectra have been measured for small protonated peptides applying photofragment-based detection schemes. These measurements allow us to investigate the role of the charge and the influence of the nature and the position of the chromophore in the peptide chain on its conformational preferences. Highly-resolved conformer-specific vibrational spectra of such species, as well as those of a double-chromophore molecule, help elucidate the mechanism of the IR-UV double-resonance methods and provide benchmarks for testing the accuracy of theoretical calculations. This thesis shows also the experimental developments that allow us to use photofragment-based detection scheme to measure highly-resolved electronic spectra of peptides of up to 17 amino acids, containing one or two chromophores, using infrared laser-assisted photofragment spectroscopy (IRLAPS). The fragmentation yield of UV pre-excited molecules increases by as much as two orders of magnitude when they further interact with the CO2 laser. This approach can be also applied in a IR-UV double resonance scheme, allowing measurements of conformer-specific infrared spectra of protonated peptides. The fragmentation channel, which is always greatly enhanced by IRMPE of UV pre-excited molecules, is the loss of the neutral, aromatic side chain via cleavage of the Cα-Cβ bond, independent of the chromophore excited in the peptide. The possible mechanisms, involving the formation of a long-lived intermediate species, that lead to the formation of the observed photofragment are discussed in this thesis work.

Official source

https://infoscience.epfl.ch/record/142377?ln=fr

À propos de ce résultat

Cette page est générée automatiquement et peut contenir des informations qui ne sont pas correctes, complètes, à jour ou pertinentes par rapport à votre recherche. Il en va de même pour toutes les autres pages de ce site. Veillez à vérifier les informations auprès des sources officielles de l'EPFL.

Concepts associés

Chargement

Publications associées

Chargement

Monia Guidi
EPFL, 2010
Thèse EPFL

Résumé

Official source

https://infoscience.epfl.ch/record/142377?ln=fr

À propos de ce résultat

Concepts associés (20)

Concepts associés

Chargement

Publications associées (62)

Publications associées

Chargement

The function of biologically active molecules depends both on their structure and on their conformational dynamics. In solution, intramolecular interaction with the solvent will influence different biological processes, i.e. protein folding. However secondary structure (helices, sheets) is heavily influenced by intramolecular forces and hence it is important to isolate biological molecules in order to study their intrinsic behavior. Moreover, gas phase studies on biomolecules provide critical information that can serve as benchmarks to test the accuracy of theoretical predictions. The main focus of this thesis is the investigation of the potential energy surfaces of biomolecules in the gas phase. Biomolecular ions are produced in the gas phase via nano-electrospray, mass-selected and guided into a cold, 22-pole ion trap where they are cooled via collisions with cold helium. A variety of double-resonance techniques based on photofragmentation detection are then applied to obtain information on the molecule stable conformations, the potential barriers separating stable conformations and their connectivity. We applied these techniques on molecules of increasing size, starting with protonated phenylalanine and proceeding to the 7amino acid peptides, Ac-Phe-(Ala)5-LysH+ and the 12-residue peptide Ac-Phe-(Ala)3-(Gly)4-(Ala)3-LysH +. As a first step, electronic spectra and conformation-specific IR-UV double resonance spectra are measured. These measurements, in combination with DFT calculations, allow the assignment of two stable conformers in the case of the protonated phenylalanine and four in the case of the seven residue peptide. In the second part of this work we focused on the conformational isomerization of these molecules by performing infrared and ultraviolet hole-filling spectroscopy in the ion trap. We demonstrated that we can induce isomerization between stable conformers of each molecule via vibrational excitation. After energy dissipation, the excited molecules are redistributed among the initially identified conformers, but no new minima were detected. In the last part of this thesis, the fractional populations and the isomerization quantum yields are determined through infrared induced population transfer spectroscopy. Protonated phenylalanine reveals conformational selectivity in its isomerization while the relaxation of the infrared excitation leads to the equilibrium distribution in the case of the 12-residue glycine-containing peptide. The steps that occur during the energy dissipation are discussed in this thesis.

EPFL2011

Chargement

Spectroscopic studies of cold, gas-phase biomolecular ions

Thomas Rizzo

While the marriage of mass spectrometry and laser spectroscopy is not new, developments over the last few years in this relationship have opened up new horizons for the spectroscopic study of biological molecules. The combination of electrospray ionization for producing large biological molecules in the gas phase together with cooled ion traps and multiple- resonance laser schemes are allowing spectroscopic investigation of individual conformations of peptides with more than a dozen amino acids. Highly resolved infrared spectra of single conformations of such species provide important benchmarks for testing the accuracy of theoretical calculations. This review presents a number of techniques employed in our laboratory and in others for measuring the spectroscopy of cold, gas-phase protonated peptides. We show examples that demonstrate the power of these techniques and evaluate their extension to still larger biological molecules.

2009

Chargement

Infrared photofragment spectroscopy of charged amino acid water clusters in the gas phase

Anthi Kamariotou

We present in this work a new technique, which combines laser photofragmentation spectroscopy with tandem mass spectrometry, for structural investigations of biomolecular ions in the gas phase. A novel apparatus was designed and built for the implementation of these studies. In this instrument closed shell ions are produced in the gas phase by electrospray ionization and stored in a hexapole ion trap prior to mass selection of a parent beam in a first quadrupole and then irradiation in an octupole ion guide using laser light. Mass analysis of the photofragmentaion products in a final stage quadrupole as a function of the wavelength generates an action spectrum. We applied this new technique to follow the microsolvation of charged amino acids in the gas phase. In these studies, the nanoelectrospray ionization source generates a distribution of water clusters of charged amino acids at various hydration levels. A particular size of cluster is selected and irradiated by infrared laser pulses resulting in the dissociation of one water molecule. Detection of the photofragmentation of this weak non-covalent bond allows us to generate the vibrational action spectrum of this particular cluster. We use a two-stage difference frequency mixing setup to produce laser light in the 2900–3800 cm-1, allowing us to probe the light-atom stretching region. IR photofragmentation spectra have been obtained for the hydrates of protonated and lithiated valine and those of protonated tryptophan. By probing the region of free and hydrogen-bonded N-H and O-H stretch vibrations and with the help of density functional theory calculations at the B3LYP/6-31++G** level, we relate spectral changes to the structure of clusters. In the study of lithiated valine water clusters, we addressed the question of zwitterion formation upon the combined effect of water and an external ion. Our data indicate the presence of the non-zwitterionic form of valine upon addition of up to four water molecules. In all of the species studied here, the hydration process is driven by solvation of the charge, and upon completion of a first shell around it, water preferentially forms a second solvation shell with no strong competition from other hydration sites on the amino acid backbone. Strikingly, similar water network structures have been observed at the highest hydration level of completely different species, probably indicating the existence of stable ordered water structures. We obtained evidence for a structrual change of the valine backbone in lithiated valine upon addition of the third water molecule, while no conformational change has been identified in the clusters of the protonated species. We have thus been able to answer questions related to the conformational preferences of the amino acid and the structuring of the water network.

EPFL2006

Chargement

Infrared photofragment spectroscopy of charged amino acid water clusters in the gas phase

Anthi Kamariotou

EPFL2006

EPFL2011