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Using high-pressure fluorescence as a method to investigate cation-π interactions in model proteins
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
(English)Manuscript (preprint) (Other academic)
Abstract [en]

This study uses High Pressure fluorescence to obtain interaction energies of weak cation-πinteractions. Since cation-π interactions are believed to contain an electrostatic component; theywould be sensitive to hydrostatic pressure. It has been established that our two model proteins,W33/K37 & W34/K38, contain cation-π interactions that are located in solvated environments.With the use of high pressure Fluorescence we were able to monitor the disruption of the cation-π interaction as well as determine the interaction energy for both solvent exposed pairs,W33/K37 & W34/K38 pairs. The weak interaction energies were found to be 0.08 ± 0.02 & 0.06± 0.02, respectively.

Keyword [en]
de novo design proteins, cation-pi interaction, high pressure fluorescence
National Category
Biological Sciences
Research subject
Biochemistry
Identifiers
URN: urn:nbn:se:su:diva-102113OAI: oai:DiVA.org:su-102113DiVA: diva2:708118
Available from: 2014-03-26 Created: 2014-03-26 Last updated: 2014-03-31
In thesis
1. Using de novo design proteins to explore tyrosine radicals and cation-π interactions
Open this publication in new window or tab >>Using de novo design proteins to explore tyrosine radicals and cation-π interactions
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Redox cofactors and amino-acid free radicals play important roles in biology. Although many of the same cofactors and amino acids that form these radicals are found across a broad range of biological systems, identical cofactors can have different reduction potentials. The local environment plays a role in defining these redox potentials. An understanding of this local-environment effect can shed more light on how redox chemistry works in nature. Our laboratory has developed a library of model proteins that are well suited to study amino-acid radicals. a3X is a de novo designed protein that is composed of 67 residues. It forms a three-helix bundle connected by two glycine loops. The radical site is located at position 32 on the central a-helix. The a3X protein is designed to be well-folded and thermodynamically stable across a broad pH range. Paper 1 describes the structural and electrochemical characterization of a3Y, a tyrosine variant of a3X. We were able to obtain a unique Faradaic response from Y32 at both low and high pH, using differential pulse voltammetry. In addition, we successfully redesigned α3Y by introducing a histidine in close proximity to Y32, creating a tyrosine/histidine pair. Our goal in creating this pair was to study proton-coupled electron transfer (PCET) in a well-structured and solvent-sequestered protein environment.  In paper 2 we illustrated the redox reversibility of Y32 and produced the first ever Pourbaix diagram for a tyrosine radical in a protein. The formal potential of the Y32-OŸ/Y32-OH redox couple was determined to be 918 ± 2 mV vs. the normal hydrogen electrode (NHE) at pH 8.40.  While at pH 5.52, the formal potential of the Y32-OŸ/Y32-OH redox couple was recorded at 1.07 V. Papers 3 and 4 utilize a3W to study cation-π interactions. In paper 3, we showed how solvation can affect the strength of these interactions by -0.9 kcal/mol. In Paper 4, we were able to monitor the disruption of the cation-π interaction with the use of high-pressure fluorescence and were able to calculate the interaction energy for a solvent exposed cation-π. The aim of the work described in this thesis was to use model proteins to study tyrosine radicals to gain a broader perspective and better understanding of the versatility of biological electron transfer and to measure cation-π interactions and how they behave in different environments.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2014. 69 p.
Keyword
tyrosine, radicals, cation-pi interactions, de novo designed proteins, biochemistry, biophysics
National Category
Biological Sciences
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-102008 (URN)978-91-7447-885-3 (ISBN)
Public defence
2014-05-09, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2014-04-14 Created: 2014-03-20 Last updated: 2014-04-14Bibliographically approved

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