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Using de novo desig...
Using de novo design proteins to explore tyrosine radicals and cation-π interactions
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- Berry, Bruce W., 1974- (författare)
- Stockholms universitet,Institutionen för biokemi och biofysik
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- Tommos, Cecilia, Research Assistant Professor (preses)
- University of Pennsylvania, Department of Biochemistry and Biophysics
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- Hofer, Anders, Associate Professor (opponent)
- Umeå university, Department of Medical Biochemistry and Biophysics
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(creator_code:org_t)
- ISBN 9789174478853
- Stockholm : Department of Biochemistry and Biophysics, Stockholm University, 2014
- Engelska 69 s.
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Abstract
Ämnesord
Stäng
- 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-O/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-O/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.
Ämnesord
- NATURVETENSKAP -- Biologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences (hsv//eng)
Nyckelord
- tyrosine
- radicals
- cation-pi interactions
- de novo designed proteins
- biochemistry
- biophysics
- Biochemistry
- biokemi
Publikations- och innehållstyp
- vet (ämneskategori)
- dok (ämneskategori)
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