Current address: Department of Biophysics, Graduate School of Science, Kyoto University, Kyoto 606-8502, Japan.
Array of Aromatic Amino Acid Side Chains Located Near the Chromophore of Photoactive Yellow Protein†
Article first published online: 28 MAR 2007
Photochemistry and Photobiology
Volume 83, Issue 2, pages 280–286, March/April 2007
How to Cite
Morishita, T., Harigai, M., Yamazaki, Y., Kamikubo, H., Kataoka, M. and Imamoto, Y. (2007), Array of Aromatic Amino Acid Side Chains Located Near the Chromophore of Photoactive Yellow Protein. Photochemistry and Photobiology, 83: 280–286. doi: 10.1562/2006-06-15-RA-929
This paper is part of the Proceedings of the 12th International Conference on Retinal Proteins held at Awaji Island, Hyogo, Japan on 4–8 June 2006.
- Issue published online: 28 MAR 2007
- Article first published online: 28 MAR 2007
- Received 15 June 2006; accepted 28 July 2006; published online 31 July 2006; DOI: 10.1562/2006-06-15-RA-929
The role of the array of aromatic amino acid side chains located close to the chromophore binding loop of photoactive yellow protein (PYP) was studied using the alanine-substitution mutagenesis. Phe92, Tyr94, Phe96 and Tyr98 were replaced with alanine (F92A, Y94A, F96A and Y98A, respectively), then these mutants were characterized by UV-visible absorption spectra, circular dichroism (CD) spectra, thermal stability and photocycle kinetics. Absorption maxima of F92A, Y94A, F96A and Y98A were 444, 442, 439 and 447 nm, respectively, different to wild type (WT) at 446 nm. Far-UV CD spectra of mutants other than F92A were different from WT, indicating that Tyr94, Phe96 and Tyr98 maintain the native secondary structure of PYP. Mid-point temperatures of thermal denaturation of F92A, Y94A and F96A, estimated by the CD signal at 222 nm, were 5–10°C lower than WT. Time constants of the photocycle estimated by flash-induced absorbance change were 0.36 s for WT and 1.4 s for Y98A, however, 100, 30 and 3000 times slower than WT for F92A, Y94A and F96A, respectively. Tyr98 is located in the loop region, whereas Phe92, Tyr94 and Phe96 are incorporated in the β4 strand, showing that aromatic amino acid residues in the β-sheet regulate the absorption spectrum, thermal stability and photocycle of PYP. Aromatic rings of Phe92, Tyr94 and Phe96 lie nearly perpendicular to the aromatic ring of Phe75 or chromophore. Possible weak hydrogen bonds between the aromatic ring hydrogen and π-electrons of these residues are discussed.