Sensitive fluorescence-based detection of magnetic field effects in photoreactions of flavins

Abstract

Magnetic field effect studies have been conducted on a variety of flavin-based radical pair systems chosen to model the magnetosensitivity of the photoinduced radical pairs found in cryptochrome flavoproteins. Cryptochromes are blue-light photoreceptor proteins which are thought to mediate avian magnetoreception, an hypothesis supported by recent in vitro observations of magnetic field-dependent reaction kinetics for a light-induced radical pair in a cryptochrome from the plant Arabidopsis thaliana. Many cryptochromes are difficult to express in large quantities or high concentrations and are easily photodegraded. Magnetic field effects are typically measured by spectroscopic detection of the transient radical (pair) concentrations. Due to its low sensitivity, single-pass transient absorption spectroscopy can be of limited use in such experiments and much recent work has involved development of other methodologies offering improved sensitivity. Here we explore the use of flavin fluorescence as the magnetosensitive probe and demonstrate the exceptional sensitivity of this technique which allows the detection of magnetic field effects in flavin samples at sub-nanomolar concentrations and in cryptochromes.

Magnetic sensitivity mediated by the Arabidopsis blue-light receptor cryptochrome occurs during flavin reoxidation in the dark

“…6 for model of the effect of the MF on the cryptochrome photocycle). It should be added that attempts in the literature to link biological magnetosensing to forward electron transfer events in cryptochrome (see, e.g., Solov'yov et al 2007;Maeda et al 2012;Mouritsen and Hore 2012;Dodson et al 2013;Biskup 2013;Evans et al 2015;Hong and Pachter 2015;Lüdemann et al 2015;Müller et al 2015;Bialas et al 2016;Kattnig et al 2016;Nohr et al 2016Nohr et al , 2017Worster et al 2016;Van der Straeten et al 2018) are not supported by the biological data neither in birds (Wiltschko et al 2016) nor (as shown in this study) in plants.…”

Detection of magnetic field effects by confocal microscopy

“…As a result, the FMN ground state population is decreased, giving rise to lower uorescence yields in the presence of a eld. 20 Finally, following baseline subtraction and the procedure outlined in the Methods section, the average %MFE can be obtained (Fig. 4b).…”

“…The % MFE of À1.25% is in excellent agreement with previous results, obtained by Evans et al in similar solution conditions. 20 Arabidopsis thaliana cryptochrome 1 (AtCry1). Fig.…”

“…18,19 Recently, however, Evans et al demonstrated that the magnetosensitivity of a radical pair generated by continuous photoexcitation may also be studied via the prompt uorescence of the photosensitiser. 20 The study focused on a avin photosensitiser and a tryptophan electron donor, a system further investigated in subsequent work by Kattnig et al using wide-eld microscopy of bulk solutions. 21 In this work, we exploit uorescence microscopy further to study photosensitive and photosensitiser-doped single crystals.…”

The Flavin–Tryptophan Dyad F10T as a Cryptochrome Model Compound: Synthesis and Photochemistry

“…The observed UV/Vis absorption and emission (fluorescence) spectra of F10T ( Figure 6) are similar to those of FMN-Trp systems (added separately in solution;F MN = flavin mononucleo- tide) reported previously. [11] While the absorption maximaa re attributable to the S 0 !S 1 (p!p*) transition of the flavin isoalloxazine moiety, the emissions pectrum corresponds to the S 1 !S 0 (p*!p)t ransition also on the flavin isoalloxazine residue. [11] These spectroscopic data showst hat the individual properties of flavin and Trpr emaininvariant in the dyad F10T.…”