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Light-induced structural changes of the LOV2 domains in various phototropins revealed by FTIR spectroscopy

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ABSTRACT

Phototropin (Phot), a blue-light photoreceptor in plants, consists of two FMN-binding domains (named LOV1 and LOV2) and a serine/threonine (Ser/Thr) kinase domain. We have investigated light-induced structural changes of LOV domains, which lead to the activation of the kinase domain, by means of light-induced difference FTIR spectroscopy. FTIR spectroscopy revealed that the reactive cysteine is protonated in both unphotolyzed and triplet-excited states, which is difficult to detect by other methods such as X-ray crystallography. In this review, we describe the light-induced structural changes of hydrogen-bonding environment of FMN chromophore and protein backbone in Adiantum neo1-LOV2 in the C=O stretching region by use of 13C-labeled samples. We also describe the comprehensive FTIR analysis of LOV2 domains among Arabidopsis phot1, phot2, and Adiantum neo1 with and without Jα helix domain.

No MeSH data available.


Difference FTIR spectra of unlabeled neo1-LOV2 (upper traces) and 12C-FMN/13C-apoprotein neo1-LOV2 (lower traces) in the 1750–1550 cm−1 region at 150 K (a) and 295 K (b). The corresponding signals are connected by dotted arrows. This figure is modified from Ref 51.
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f4-7_89: Difference FTIR spectra of unlabeled neo1-LOV2 (upper traces) and 12C-FMN/13C-apoprotein neo1-LOV2 (lower traces) in the 1750–1550 cm−1 region at 150 K (a) and 295 K (b). The corresponding signals are connected by dotted arrows. This figure is modified from Ref 51.

Mentions: Both C4=O and C2=O stretching vibrations shift to higher frequencies upon the formation of S390 (Fig. 3), which was essentially temperature-independent51. This shows that the hydrogen bonds of the C=O groups are weakened by adduct formation, which presumably relocates the FMN chromophore apart from their hydrogen-bonding donors. The conformational switch of Gln1029 may also contribute to the frequency change in S39050. It should be noted, however, that there is a certain temperature-dependence for the C4=O and C2=O stretching vibrations in the S390 state but in the unphotolyzed state. The frequencies of the C4=O stretch in S390 are 1723 and 1730 cm−1 at 150 and 295 K, respectively, whereas those of the C2=O stretch in S390 are 1687 and 1683 cm−1 at 150 and 295 K, respectively (see below; Fig. 4). These frequency shifts possibly originate from temperature-dependent chromophore-protein interaction changes.


Light-induced structural changes of the LOV2 domains in various phototropins revealed by FTIR spectroscopy
Difference FTIR spectra of unlabeled neo1-LOV2 (upper traces) and 12C-FMN/13C-apoprotein neo1-LOV2 (lower traces) in the 1750–1550 cm−1 region at 150 K (a) and 295 K (b). The corresponding signals are connected by dotted arrows. This figure is modified from Ref 51.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC5036776&req=5

f4-7_89: Difference FTIR spectra of unlabeled neo1-LOV2 (upper traces) and 12C-FMN/13C-apoprotein neo1-LOV2 (lower traces) in the 1750–1550 cm−1 region at 150 K (a) and 295 K (b). The corresponding signals are connected by dotted arrows. This figure is modified from Ref 51.
Mentions: Both C4=O and C2=O stretching vibrations shift to higher frequencies upon the formation of S390 (Fig. 3), which was essentially temperature-independent51. This shows that the hydrogen bonds of the C=O groups are weakened by adduct formation, which presumably relocates the FMN chromophore apart from their hydrogen-bonding donors. The conformational switch of Gln1029 may also contribute to the frequency change in S39050. It should be noted, however, that there is a certain temperature-dependence for the C4=O and C2=O stretching vibrations in the S390 state but in the unphotolyzed state. The frequencies of the C4=O stretch in S390 are 1723 and 1730 cm−1 at 150 and 295 K, respectively, whereas those of the C2=O stretch in S390 are 1687 and 1683 cm−1 at 150 and 295 K, respectively (see below; Fig. 4). These frequency shifts possibly originate from temperature-dependent chromophore-protein interaction changes.

View Article: PubMed Central - PubMed

ABSTRACT

Phototropin (Phot), a blue-light photoreceptor in plants, consists of two FMN-binding domains (named LOV1 and LOV2) and a serine/threonine (Ser/Thr) kinase domain. We have investigated light-induced structural changes of LOV domains, which lead to the activation of the kinase domain, by means of light-induced difference FTIR spectroscopy. FTIR spectroscopy revealed that the reactive cysteine is protonated in both unphotolyzed and triplet-excited states, which is difficult to detect by other methods such as X-ray crystallography. In this review, we describe the light-induced structural changes of hydrogen-bonding environment of FMN chromophore and protein backbone in Adiantum neo1-LOV2 in the C=O stretching region by use of 13C-labeled samples. We also describe the comprehensive FTIR analysis of LOV2 domains among Arabidopsis phot1, phot2, and Adiantum neo1 with and without Jα helix domain.

No MeSH data available.