Tripping the light fantastic: blue-light photoreceptors as examples of environmentally modulated protein-protein interactions

Abstract

Blue-light photoreceptors play a pivotal role in detecting the quality and quantity of light in the environment, controlling a wide range of biological responses. Several families of blue-light photoreceptors have been characterized in detail using biophysics and biochemistry, beginning with photon absorption, through intervening signal transduction, to regulation of biological activities. Here we review the light oxygen voltage, cryptochrome, and sensors of blue light using FAD families, three different groups of proteins that offer distinctly different modes of photochemical activation and signal transduction yet play similar roles in a vast array of biological responses. We cover mechanisms of light activation and propagation of conformational responses that modulate protein-protein interactions involved in biological signaling. Discovery and characterization of these processes in natural proteins are now allowing the design of photoregulatable engineered proteins, facilitating the generation of novel reagents for biochemical and cell biological research.

Figure 1. Representative structures of the four families of blue light photoreceptors

A) Arabidopsis CRY1 (1U3C) (42), with the two domains (helical in yellow, αβ in blue) linked by an extended linker. The FAD chromophore is depicted in magenta. B) AsLOV2 (2VOU) (76) contains a typical PAS domain fold with the PAS core (blue) β-sheet flanked by Ncap (yellow) and Ccap (red) elements. C) Structure of AppA 1–125 (2IYJ) (109). The dynamic β4-β5 loop is depicted in blue. D) PYP (2PHY) (22) is also comprised of a typical PAS fold with covalently-attached 4-hydroxycinnamic acid chromophore (green). Similar to AsLOV2 an Ncap is present (yellow).

Figure 2. Flavin chemistry

FAD or (FMN) can undergo a series of electron and proton transfers to form FAD (oxidized), FAD⁻ (anionic semiquinone), FADH (neutral semiquinone) and FADH⁻ (hydroquinone). Flavin species often perform chemistry at the C4a and N5 positions, including the generation of covalent adducts to both locations.

Figure 3. Schematic of cryptochrome activation and protein:protein interactions

The CRY CCT can either act as a directly repressive element inhibiting interactions at the PHR domain (top Drosophila model) or as a recognition element (bottom Arabidopsis model). In Drosophila, dCRY has multiple recognition partners, while the active state of Arabidopsis CRY is a dimer that interacts with COP1.

Figure 4. Models of signal propagation in LOV domains

Both AsLOV2 (A, B) and VVD (C) initiate signal propagation from rotation of a conserved Gln in the Iβ strand (Q513 in AsLOV2, Q182 in VVD). Rotation induces interactions with residues at the beginning of the adjacent Aβ strand, which induces alterations of H-bonds between Asp414 (AsLOV2) or induces rotation of Cys71 (VVD). Direct observation of movement of the Ncap was seen in VVD light-state crystals.

Figure 5. Schematic of LOV domain signaling

LOV domains sequester signaling elements via Ccap or Ncap elements. In VVD, conformational changes at the Ncap induce dimerization involving Ncap elements. In AsLOV2, Ccap elements lock the signaling domain (kinase blue) in an inactive conformation. Changes in Jα docking relieve inhibition.

Figure 6. BLUF domain activation and signaling

A) Mechanism of BLUF activation. Light induces radical transfer from Tyr21 to FAD, which induces tautomerization of Gln63. B) Comparison of Trp_in (Blue) and Trp_out (yellow) conformations. Movement of Trp results in alteration of the β5 loop and adjoining helix. These perturb the conformation of Ccap regions bound on the opposite side of the central β-sheet (grey), altering effector function.

Figure 7. Structure of a photoactivatable LOV-Rac fusion

The dark state inhibits the activity of Rac (grey) via interactions between AsLOV2 (blue and red) and Rac (2WKP) (150). Light-induced unfolding of the Jα helix (red) can relieve inhibition, presumably by allowing freedom between the two domains.