R. van Grondelle
- Conformational heterogeneity and propagation of structural changes in the LOV2/Jα domain from Avena sativa phototropin 1 as recorded by temperature-dependent FTIR spectroscopy
- Biophysical Journal
- Volume | Issue number
- 97 | 1
- Pages (from-to)
- Document type
- Faculty of Science (FNWI)
- Swammerdam Institute for Life Sciences (SILS)
Phototropins control phototropism, chloroplast movement, stomatal opening, and leaf expansion in plants. Phototropin 1 (phot1) is composed of a kinase domain linked to two blue light-sensing domains, LOV2 and LOV1, which bind flavin mononucleotide. Disruption of the interaction between the LOV2 domain and a helical segment named Jα, joining LOV to the kinase domain, induces the subsequent kinase activity of phototropin 1 and further-downstream signal transduction. Here we study the effects of temperature and hydration on the light-triggered signal propagation in the phot1 LOV2 domain of Avena sativa (AsLOV2/Jα), using Fourier transform infrared spectroscopy to unravel part of the molecular mechanism of phototropin 1. We report that AsLOV2/Jα shows an intense signal in the amide I and II regions, arising mainly from β-sheet changes and the unbinding of the Jα helix from the Per-ARNT-Sim core and its subsequent partial unfolding. Importantly, these structural changes only occur under conditions of full hydration and at temperatures above 280 K. We characterized a newly isolated low-hydration intermediate that shows a downshift of high-frequency amide I signals and that possibly corresponds to loop tightening, without large β-sheet or Jα structural changes. In addition, we report a heterogeneity in AsLOV2/Jα involving two different C(4)=O conformer populations, coexisting in the dark state and characterized by C(4)=O carbonyl frequencies at 1712 cm−1 and 1694 cm−1 that are attributable to a single H-bond and two H-bonds at this site, respectively. Such conformers display slightly shifted absorption spectra and cause a splitting of the 475-nm band in the ultraviolet/visible spectra of LOV domains at low temperature.
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