Photocontrol of protein-protein and protein-nucleic acid interactions

Kneissl, Sabine 2009. Photocontrol of protein-protein and protein-nucleic acid interactions. PhD Thesis, Cardiff University.

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Abstract

Proteins often depend on a-helices for binding to other biomacromolecules. Reversible control of a-helix stability was accomplished in previous studies by incorporating a photoisomerisable azobenzene cross-linker into peptides, subsequently enabling the optical control of DNA-protein interactions. This approach was extended in this study to include protein-protein and protein-RNA interactions. One of the primary regulatory components in apoptosis signalling is the antiapoptotic protein Bcl-xL which interacts with the a-helical BH3 domain of the Bak protein. The Rev/RRE interaction is crucially involved in the life cycle of Human Immunodeficiency Virus. These interactions were targeted by designing peptides based on the BH3 domain of Bak and on the RNA-binding domain of Rev these peptides are activated by external light pulses after the incorporation of the cross-linker. The ability to control cross-linker conformation and hence peptide secondary structure was demonstrated by CD and UV/Vis spectroscopy. The binding to the target structure and complex disruption was determined in the dark-adapted and irradiated states using fluorescence based assays. Structural studies using NMR spectroscopy demonstrated that the alkylated peptides bind to the same part of the target molecule as the wild-type peptide, regardless of their structure. Moreover, one of the BH3 domain-based peptides and the light-controllable transcription factor PhotoMyoD were modified with protein transduction domains to enable future in vivo studies. Overall, this work opens the possibility to interfere reversibly and specifically with protein-protein and protein-RNA interactions and to study and modulate cellular function by optical control.

Item Type:	Thesis (PhD)
Status:	Unpublished
Schools:	Chemistry
Subjects:	Q Science > QD Chemistry
ISBN:	9781303214516
Date of First Compliant Deposit:	30 March 2016
Last Modified:	25 Oct 2017 14:35
URI:	https://orca.cardiff.ac.uk/id/eprint/54835

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