Engineering of Halorhodospira halophila photoactive yellow protein.
Contributors: University of Toronto.  Dept. of Chemistry.
Published: 2005
Language: eng
Number of pages: 70
Description: Halorhodospira halophila photoactive yellow protein (PYP) is a promising candidate to act as a photoswitching domain in engineered proteins. Absorption of a photon of wavelength 446 nm triggers trans  to cis isomerization of its 4-hydroxycinnamic acid chromophore leading to large structural perturbations in the protein's N-terminal region. In the dark, a slower cis to trans reisomerization of the chromophore occurs leading to restoration of the protein's native fold. Thus, fusion of proteins to PYP's N-terminus may enable modulation of the activity of the attached molecule by making use of the PYP photocycle.Site-directed mutagenesis was used to mutate the initiator ATG codon of PYP to TGC. This facilitates expression of PYP with an N-terminal cysteine allowing for N-terminal fusions to PYP using native chemical ligation. Our approach has been to use the amino thioacetic acid S-benzyl ester of 5-carboxy-tetramethylrhodamine (TMR),6, fluorophore, as well as S-Peptide for N-terminal attachment to PYP.
Notes: Source: Masters Abstracts International, Volume: 44-02, page: 0906.

Advisor: G. Andrew Woolley

Thesis (M.Sc.)--University of Toronto, 2005.

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