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Time-resolved infrared spectroscopy of photoswitchable thioxopeptides


Bregy, H. Time-resolved infrared spectroscopy of photoswitchable thioxopeptides. 2009, University of Zurich, Faculty of Science.

Abstract

Proteins are important biomolecules. Knowledge of their conformations and conformational dynamics is essential to understand their biological functionality. While there exist established methods to investigate the structure of a protein, methods to explore the dynamics like protein folding, allosteric regulations and catalysis are relatively new and still in development. Processes which happen on a millisecond or slower time scale can be eciently studied by NMR spectroscopy. To investigate faster dynamical processes, other spectroscopic methods are needed. In this work, time-resolved infrared spectroscopy on small peptides reveals information about dynamics on the picosecond time scale. Twodimensional infrared spectroscopy (2D-IR) obtains structural information like coupling and relative dipole orientations between molecular groups in a protein/peptide in equilibrium. This method can also be applied to a system being not in equilibrium, e.g a peptide undergoing conformational change (transient 2D-IR). Transient (2D-)IR experiments require a trigger to induce structural changes at a well dened time. This trigger can be e.g. a temperature-jump or a molecular photoswitch in the peptide. In this work, the substitution of the carbonyl oxygen in one peptide bond by a sulfur atom is used to create a photoswitch in the peptide. The resulting thioamide bond then can be excited selectively by ultraviolet light ( ! excitation red-shifted in regard with an amide bond). This moiety then isomerizes and forces the peptide to change its structure. To investigate the functioning of the thioamide bond as photoswitch, N-Methylthioacetamide has been investigated by dierent methods revealing trans ! cis isomerization with a high yield which is completed within one nanosecond. In this work, a thioamide containing peptide, Boc-Ala-Pro- (SC-NH)-Aib-Ala-OMe, is synthesized. This thioxopeptide has well-resolved bands in the amide I region which makes the interpretation of transient spectra easier. Furthermore, it contains a -Pro-Aib- amino acid sequence which is known to form stable -turn structure, characterized by a i ! i+3 hydrogen-bond. The breaking of this hydrogen-bond, induced by the photoswitch, is investigated by transient infrared spectroscopy (UV-pump/IR-probe spectroscopy). The dynamics is compared to other - Aib-containing thioxopeptides which tend to form -turns as well. All thioxopeptides show very similar photoisomerization dynamics, while there are signicant dierences for the relaxation in the thermal ground state. Furthermore, structural properties are revealed from the time-resolved measurements, which show very rigid structure in the inner part of all the thioxopeptides (- (SC-NH)-Aib-), while the tails are exible. Finally, the investigation on the i ! i+3 hydrogen-bond breaking of thioxopeptide Boc-Ala-Pro- (SC-NH)- Aib-Ala-OMe is extended to transient 2D-IR spectroscopy. For detailed interpretation of the spectra, they are compared to additionally performed computer simulations.

Proteins are important biomolecules. Knowledge of their conformations and conformational dynamics is essential to understand their biological functionality. While there exist established methods to investigate the structure of a protein, methods to explore the dynamics like protein folding, allosteric regulations and catalysis are relatively new and still in development. Processes which happen on a millisecond or slower time scale can be eciently studied by NMR spectroscopy. To investigate faster dynamical processes, other spectroscopic methods are needed. In this work, time-resolved infrared spectroscopy on small peptides reveals information about dynamics on the picosecond time scale. Twodimensional infrared spectroscopy (2D-IR) obtains structural information like coupling and relative dipole orientations between molecular groups in a protein/peptide in equilibrium. This method can also be applied to a system being not in equilibrium, e.g a peptide undergoing conformational change (transient 2D-IR). Transient (2D-)IR experiments require a trigger to induce structural changes at a well dened time. This trigger can be e.g. a temperature-jump or a molecular photoswitch in the peptide. In this work, the substitution of the carbonyl oxygen in one peptide bond by a sulfur atom is used to create a photoswitch in the peptide. The resulting thioamide bond then can be excited selectively by ultraviolet light ( ! excitation red-shifted in regard with an amide bond). This moiety then isomerizes and forces the peptide to change its structure. To investigate the functioning of the thioamide bond as photoswitch, N-Methylthioacetamide has been investigated by dierent methods revealing trans ! cis isomerization with a high yield which is completed within one nanosecond. In this work, a thioamide containing peptide, Boc-Ala-Pro- (SC-NH)-Aib-Ala-OMe, is synthesized. This thioxopeptide has well-resolved bands in the amide I region which makes the interpretation of transient spectra easier. Furthermore, it contains a -Pro-Aib- amino acid sequence which is known to form stable -turn structure, characterized by a i ! i+3 hydrogen-bond. The breaking of this hydrogen-bond, induced by the photoswitch, is investigated by transient infrared spectroscopy (UV-pump/IR-probe spectroscopy). The dynamics is compared to other - Aib-containing thioxopeptides which tend to form -turns as well. All thioxopeptides show very similar photoisomerization dynamics, while there are signicant dierences for the relaxation in the thermal ground state. Furthermore, structural properties are revealed from the time-resolved measurements, which show very rigid structure in the inner part of all the thioxopeptides (- (SC-NH)-Aib-), while the tails are exible. Finally, the investigation on the i ! i+3 hydrogen-bond breaking of thioxopeptide Boc-Ala-Pro- (SC-NH)- Aib-Ala-OMe is extended to transient 2D-IR spectroscopy. For detailed interpretation of the spectra, they are compared to additionally performed computer simulations.

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Additional indexing

Item Type:Dissertation
Referees:Hamm P, Seeger S, Hutter J
Communities & Collections:07 Faculty of Science > Department of Chemistry
Dewey Decimal Classification:540 Chemistry
Language:English
Date:2009
Deposited On:18 Feb 2010 18:32
Last Modified:05 Apr 2016 13:55
Number of Pages:111
Related URLs:http://opac.nebis.ch/F/?local_base=NEBIS&con_lng=GER&func=find-b&find_code=SYS&request=005873297
Permanent URL: http://doi.org/10.5167/uzh-30796

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