Conformational Dynamics of Phototropin 2 LOV2 Domain with the Linker upon Photoexcitation

Abstract

Conformational dynamics of LOV2 domain of phototropin, a plant blue light photoreceptor, is studied by the pulsed laser induced transient grating (TG) technique. The TG signal of LOV2 without the linker part to the kinase domain exhibits the thermal grating signal due to the heat releasing from the excited state and a weak population grating by the adduct formation. The diffusion coefficients of the adduct product after forming the chemical bond between the chromophore and Cys residue are found to be slightly smaller than that of the reactant, which implies that the core shrinks slightly on the adduct formation. After that change, no significant conformational change was observed. On the other hand, the signal of LOV2 with the linker part to the kinase domain clearly shows very different diffusion coefficients between the original and the adduct species. The large difference indicates significant global conformational change of the protein moiety upon the adduct formation. More interestingly, the diffusion coefficient is found to be time-dependent in the observation time range. The dynamics representing the global conformational change is a clear indication of a spectral silent intermediate between the excited triplet state and the signaling product. From the temporal profile analysis of the signal, the rate of the conformational change is determined to be 2 ms.

Photosensitivity of Kinase Activation by Blue Light Involves the Lifetime of a Cysteinyl-Flavin Adduct Intermediate, S390, in the Photoreaction Cycle of the LOV2 Domain in Phototropin, a Plant Blue Light Receptor

“…However, UV-visible absorption reflects the electronic environments around the isoalloxazine ring of the FMN chromophore and cannot sense the conformational changes in the protein moiety. We therefore proposed denoting these photointermediates with a UVvisible absorption spectrum of S390 and conformational changes in the protein moiety detected by NMR (15), SAXS (16), FTIR (17,18), or transient grating (19) as S390II (37). In the present investigation, the phot proteins consisting of LOV2-linker-kinase also exhibited production of a photointermediate (S390) similar to that of LOV cores accompanied by kinase activation.…”

“…Light-induced conformational changes in the linker region were detected in At phot1 with small angle x-ray scattering (SAXS) (16), FTIR spectroscopy (17,18), and transient grating (19). Recently, light-induced movement of the LOV2 domain relative to the kinase domain in At phot2 was visualized by SAXS (20).…”

Light-Induced Conformational Change and Transient Dissociation Reaction of the BLUF Photoreceptor Synechocystis PixD (Slr1694)

“…6,[23][24][25][26] The time profile represents the reaction kinetics as well as the thermal diffusion and molecular diffusion processes. Figure 1 shows a typical TG signal after photoexcitation of PixD in the buffer solution at a grating wavenumber q 2 = 4.0 × 10 11 m − 2 and a laser power of 0.2 mJ cm − 2 .…”

Mutational Analysis of Phototropin 1 Provides Insights into the Mechanism Underlying LOV2 Signal Transmission

“…5). FTIR spectroscopy (26,27,36,37), CD spectroscopy (38), small angle x-ray scattering (19), and pulsed laser-induced transient gating methods (39) have shown that irradiation of LOV2-J␣ protein fragments derived from several different phototropin sources results in significant structural changes at the surface of the LOV-core. In particular, FTIR spectroscopy has indicated that light-induced formation of LOV2 390 produces progressive changes that involve regions of loops, ␣-helices, and ␤-sheets (26,27).…”