Detecting a New Source for Photochemically Induced Dynamic Nuclear Polarization in the LOV2 Domain of Phototropin by Magnetic-Field Dependent 13C NMR Spectroscopy
 

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Detecting a New Source for Photochemically Induced Dynamic Nuclear Polarization in the LOV2 Domain of Phototropin by Magnetic-Field Dependent 13C NMR Spectroscopy


Kothe, G., et al., Detecting a New Source for Photochemically Induced Dynamic Nuclear Polarization in the LOV2 Domain of Phototropin by Magnetic-Field Dependent 13C NMR Spectroscopy. The Journal of Physical Chemistry B, 2014. 118(40): p. 11622-11632.


http://dx.doi.org/10.1021/jp507134y


<div style="text-align: justify;">Phototropin is a flavin mononucleotide (FMN) containing blue-light receptor, which regulates, governed by its two LOV domains, the phototropic response of higher plants. Upon photoexcitation, the FMN cofactor triplet state, 3F, reacts with a nearby cysteine to form a covalent adduct. Cysteine-to-alanine mutants of LOV domains instead generate a flavin radical upon illumination. Here, we explore the formation of photochemically induced dynamic nuclear polarization (CIDNP) in LOV2-C450A of Avena sativa phototropin and demonstrate that photo-CIDNP observed in solution 13C NMR spectra can reliably be interpreted in terms of solid-state mechanisms including a novel triplet mechanism. To minimize cross-polarization, which transfers light-induced magnetization to adjacent 13C nuclei, our experiments were performed on proteins reconstituted with specifically 13C-labeled flavins. Two potential sources for photo-CIDNP can be identified: The photogenerated triplet state, 3F, and the triplet radical pair 3(F??W+?), formed by electron abstraction of 3F from tryptophan W491. To separate the two contributions, photo-CIDNP studies were performed at four different magnetic fields ranging from 4.7 to 11.8 T. Analysis revealed that, at fields