Abstract
The AppA BLUF (blue light sensing using FAD) domain from Rhodobacter sphaeroides serves as a blue light-sensing photoreceptor. The charge separation process between Tyr-21 and flavin plays an important role in the light signaling state by transforming the dark state conformation to the light state one. By solving the linearized Poisson-Boltzmann equation, I calculated E(m) for Tyr-21, flavin, and redox-active Trp-104 and revealed the electron transfer (ET) driving energy. Rotation of the Gln-63 side chain that converts protein conformation from the dark state to the light state is responsible for the decrease of 150 mV in E(m) for Tyr-21, leading to the significantly larger ET driving energy in the light state conformation. The pK(a) values of protonation for flavin anions are essentially the same in both dark and light state crystal structures. In contrast to the ET via Tyr-21, formation of the W state results in generation of only the dark state conformation (even if the initial conformation is in the light state); this could explain why Trp-104-mediated ET deactivates the light-sensing yield and why the activity of W104A mutant is similar to that of the light-adapted native BLUF.
Highlights
Electron donors in the charge separation process, where Tyr-21ϩ. is the functionally relevant charge state, whereas formation of Trp-104ϩ. does not contribute to light sensing of AppA BLUF (4)
B Conformation where Trp-104 is in the FMN binding pocket. c See Ref. 7. d The –NH2 and –CO groups of Gln-63 were rotated by 180° along the C␥-C␦ axis, and their positions were energetically optimized with CHARMM (18). e Em(W/Wϩ. ) in the BLUF domain. f Em(W/Wϩ. ) in water (26). g Shift in Em(W/Wϩ. ) from water to protein. h Influence of uncharged protein dielectric volume. i Influence of the FMN 5Ј-phosphate group
Demonstrated that (i) the W104A mutant is insensitive to blue light and (ii) its activity is similar to that of the light-adapted native BLUF
Summary
Em(Y/Yϩ. ), I obtained the driving energy (⌬G) of the ET between FMN and Y to be Ϫ156 and Ϫ317 meV for the dark and light state structures in the charge-separated [Yϩ. -W-FMN. ] state, respectively (Table 1). ), I obtained the driving energy (⌬G) of the ET between FMN and Y to be Ϫ156 and Ϫ317 meV for the dark and light state structures in the charge-separated [Yϩ. Since the light state structure is energetically much favorable for the ET than the dark state structure in the present study, the charge separation process via Tyr-21 Ϫ350 ND a Dark (light) state structure where the –NH2 (–CO) group of the Gln-63 side chain is a hydrogen-bonding partner of Tyr-21. ). and not Driving Force of Side Chain Rotation of Gln-63—Regardless of the difference in the hydrogen bond pattern with regard to the Gln-63 and FMN pair, the calculated pKa(N5) values of FMN are essentially the same in both dark and light state structures (pKa(N5) 14.0 and 13.9 in ϭ 12.2 and 12.3 in the [Y-W-FMN] state and the [Yϩ. I(nYfl؉.u-eWn-cFeMoNf t.h)ecpornoftoerimnaetniovniroinnmmeVnt on Em(Y/Y؉. ) for theY-W-FMN
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
