Chemical amplification of magnetic field effects relevant to avian magnetoreception

Journal name:
Nature Chemistry
Volume:
8,
Pages:
384–391
Year published:
DOI:
doi:10.1038/nchem.2447
Received
Accepted
Published online

Abstract

Magnetic fields as weak as the Earth's can change the yields of radical pair reactions even though the energies involved are orders of magnitude smaller than the thermal energy, kBT, at room temperature. Proposed as the source of the light-dependent magnetic compass in migratory birds, the radical pair mechanism is thought to operate in cryptochrome flavoproteins in the retina. Here we demonstrate that the primary magnetic field effect on flavin photoreactions can be amplified chemically by slow radical termination reactions under conditions of continuous photoexcitation. The nature and origin of the amplification are revealed by studies of the intermolecular flavin–tryptophan and flavin–ascorbic acid photocycles and the closely related intramolecular flavin–tryptophan radical pair in cryptochrome. Amplification factors of up to 5.6 were observed for magnetic fields weaker than 1 mT. Substantial chemical amplification could have a significant impact on the viability of a cryptochrome-based magnetic compass sensor.

At a glance

Figures

  1. Intermolecular radical pair reactions of flavins.
    Figure 1: Intermolecular radical pair reactions of flavins.

    Reaction scheme for the intermolecular photochemical reaction of a flavin (F) with an electron donor (D). The radicals, F•− and D•+, are either charged as shown or (de)protonated depending on the pH. The slow reaction steps labelled with the rate constants kD and kF are responsible for the amplification of the MFE. The other reaction steps are as follows: photoexcitation of the ground-state flavin (F) to its first excited singlet state (1F*); fluorescence of 1F*; intersystem crossing from 1F* to the excited triplet state of the flavin (3F*); spin-conserving electron transfer from the donor D to 3F* to form the triplet state of the radical pair, 3(F•− D•+); coherent interconversion of the triplet and singlet, 1(F•− D•+), radical pair states driven by hyperfine and Zeeman magnetic interactions (curly arrows); spin-allowed reverse electron transfer within the singlet radical pair, which regenerates the ground state of both reactants; diffusive separation to form free radicals, which subsequently re-encounter to form radical pairs. Singlet–triplet interconversion becomes less efficient in a magnetic field stronger than the hyperfine interactions because only the T0 triplet substate is able to mix with the singlet, whereas T+1 and T−1, as well as T0, mix with the singlet when the external magnetic field is weaker than the hyperfine interactions.

  2. Amplified MFEs in intermolecular reactions of flavins.
    Figure 2: Amplified MFEs in intermolecular reactions of flavins.

    a, Fluorescence intensity of an aqueous solution (pH 4.0) of FMN (10 µM) and lysozyme (0.5 mM) as a function of the time after the onset of continuous illumination (470 nm) with an applied magnetic field switched between 0 and 27 mT every 1.68 seconds. Modulation of the fluorescence intensity, shown expanded in the inset, occurs in synchrony with the magnetic field switching. b, The average of the responses in a to one cycle of the magnetic field shows a prompt MFE (χp) and a slowly established delayed MFE (χd). c, Dependence of the MFE on the magnetic field strength for different delay times between switching the field on and measuring the fluorescence. The dashed lines are Lorentzian fits to the data used to determine the B1/2 values. d, As a, but for a solution (pH 6.2) of FMN (10 µM) and Trp (1.0 mM). The magnetic field was switched between 0 and 27 mT every 0.56 seconds. e, Average of the responses in d to one cycle of the magnetic field. In contrast to b, the delayed MFE (χd) is smaller than the prompt effect (χp). f, As e, but with the pH reduced to 2.7 (citric acid buffer) (blue) and with 50 µM TCEP (pH 6.2) (orange), which shows that the amplification factor can be changed by altering the rate constants kD and kF. The data in the field-on and field-off periods in a, b, d and e are shown in green and blue, respectively. The timing of the magnetic field steps is shown in grey at the top of b, e and f (B0 is the magnetic field strength). In b and e, the slow background decay of the fluorescence (visible in a and d) has been subtracted.

  3. Characteristics of the amplified MFE.
    Figure 3: Characteristics of the amplified MFE.

    a, Response of the concentration of the flavin excited state, 1F*, to sudden on/off switching of an external magnetic field (B0). Left, kF = kD (E = 1); centre, kD/kF < 1 (E < 1); right, kD/kF > 1 (E > 1). Depending on the ratio of their rate constants, kD/kF, the radical termination reactions lead to a slow kinetic phase that enhances or attenuates the prompt MFE. b, Contour plot of the amplification factor, E, as a function of the slow radical termination rate constants kF and kD. E depends principally on the ratio kD/kF; the largest amplification factors occur when kD >> kF. The simulations were performed for an initial triplet radical pair and a strong magnetic field. The values of the other rate constants used in the simulation are given in Supplementary Section C.

  4. Amplified MFEs in the low-field region.
    Figure 4: Amplified MFEs in the low-field region.

    a, Dependence of the MFE on the magnetic field strength for an aqueous solution (pH 3.8) of FMN (10 µM) and ascorbic acid (0.5 mM) with a 50 ms delay time between switching the field on and measuring the fluorescence. The positive MFE at fields below 1.4 mT is known as the low field effect; it is particularly prominent in this case because of the small hyperfine interactions in the ascorbyl radical. b, Average of the responses to one cycle of a magnetic field switched between 0 and 27 mT every 1.12 seconds. c, As b, but for a 0.7 mT magnetic field (in the region of the low field effect). The prompt and delayed components have opposite phases in b and c because of the biphasic form of the MFE (shown in a). The black lines in b and c are fits to single exponentials.

  5. Intramolecular radical pair reactions of cryptochromes.
    Figure 5: Intramolecular radical pair reactions of cryptochromes.

    Reaction scheme for the intramolecular photochemistry of the FAD (F) and the terminal residue of the Trp triad in cryptochrome as the electron donor (D). RP2 is a FAD–Trp radical pair in which the flavin radical (F•−) is protonated or the Trp radical (D•+) is deprotonated or both; it is not magnetically sensitive. The slow steps labelled with the rate constants kD and kF are responsible for the amplification of the MFE. As shown, we assume that the flavin radical may be oxidized before or after the Trp radical is reduced.

  6. Amplified MFEs in a cryptochrome.
    Figure 6: Amplified MFEs in a cryptochrome.

    a, Changes in the fluorescence spectrum of an aqueous solution of AtCry1 (about 100 µM, 10 mM Tris, pH 7.4, 150 mM NaCl, 30% glycerol v/v) induced by a magnetic field. Spectra (smoothed using a 25 nm moving average filter) are shown for different delay times, T, after switching on a 12.2 mT magnetic field, having subtracted the zero-field spectrum. The MFE becomes dramatically stronger as T is increased from 0.5 to 3.5 seconds. The black trace is the fluorescence spectrum measured in the absence of a magnetic field and scaled to match the maximum of the difference spectrum at T = 3.5 seconds (red trace). b, Average of the responses of an aqueous solution of AtCry1 (about 100 µM, pH 7.4) to one cycle of a magnetic field switched between 0 and 27 mT every 4.2 seconds. The enhanced MFE grows over a period of several seconds as a result of the slow radical termination reactions (kD > kF). c, As b but with the addition of TCEP (50 µM). An accelerated reduction of the Trp•+ radical by the TCEP boosts the MFE (kD >> kF) and causes it to grow more rapidly. In b and c the black lines are fits of the data to single exponentials assuming symmetric responses to switching the field on and off.

  7. Simulations of changes in radical concentrations induced by a magnetic field.
    Figure 7: Simulations of changes in radical concentrations induced by a magnetic field.

    a, Density plot of the absolute prompt change in the total concentration of flavin radicals produced by an applied magnetic field, plotted as a function of the radical termination rate constants kF and kD. b, As a, but showing the absolute steady-state change induced by a magnetic field. The scale bar refers to both plots. The amplification factor in the centre of the yellow region of b, where the absolute steady-state change is largest, has a value of 33.

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Author information

  1. These authors contributed equally

    • Daniel R. Kattnig &
    • Emrys W. Evans
  2. Present address: Molecular Medicine, National Heart & Lung Institute, Imperial College London, London SW7 2AZ, UK

    • Charlotte A. Dodson

Affiliations

  1. Department of Chemistry, University of Oxford, Physical & Theoretical Chemistry Laboratory, Oxford OX1 3QZ, UK

    • Daniel R. Kattnig,
    • Stuart R. Mackenzie &
    • P. J. Hore
  2. Department of Chemistry, University of Oxford, Inorganic Chemistry Laboratory, Oxford OX1 3QR, UK

    • Emrys W. Evans,
    • Victoire Déjean &
    • Christiane R. Timmel
  3. Department of Chemistry, University of Oxford, Chemistry Research Laboratory, Oxford OX1 3TA, UK

    • Charlotte A. Dodson &
    • Mark I. Wallace

Contributions

D.R.K. and E.W.E. contributed equally to this work. D.R.K., E.W.E. and V.D. designed and performed the experiments. D.R.K. and E.W.E. analysed the data. C.A.D. advised on the production of the AtCry1 samples. M.I.W. helped oversee the fluorescence microscopy experiments. C.R.T., S.R.M. and P.J.H. coordinated the study. P.J.H., D.R.K. and E.W.E. wrote the paper. All the authors discussed the results and commented on the manuscript.

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The authors declare no competing financial interests.

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