Abstract
Recently, BLUF proteins were found as flavin-binding blue light sensors in some bacteria and eukaryotes. BLUF domain shows red-shifted absorption spectrum when illuminated by blue light. The conserved Tyr-21 of AppA was a critical amino acid residue for this photocycle in Rhodobacter sphaeroides. Previously, we reported that BLUF protein Tll0078 of a cyanobacterium Thermosynechococcus elongates BP-1 showed such photocycle. Here, we generated the site-directed mutated Tll0078 at the conserved Tyr-8. Tll0078_Y8A protein, where Tyr-8 was replaced with Ala, was overexpressed and purified from Escherichia coli. Purified Tll0078_Y8A bound flavin, but did not show red-shifted absorption spectrum upon illumination. On the other hand, the absorbance around 370 ~ 470 nm bleached when illuminated for longer time and slowly recovered when in the dark. This may suggest light dependent reduction of flavin. We will discuss about mechanism of photocycle and structure of near flavin of BLUF domain.
