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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.


(a) Schematic illustration of the LOV2 constructs of phototropin presented in this review. For the reconstitution of Adiantum neo1-LOV2 (*Ac neo1), we used the P905–P1087 construct containing the CBP tag at the N-terminus. For the comparison of the effect of Jα helix, we used the constructs encoding the same region of the LOV2-core and LOV2-Jα for Arabidopsis (At) phot1 and phot2 and Adiantum (Ac) neo1, where the GST tag at the N-terminus is digested with thrombin. (b) Amino acid sequences of the C-terminal side of LOV2-core and LOV2-Jα. This figure is modified from Ref 53.
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f2-7_89: (a) Schematic illustration of the LOV2 constructs of phototropin presented in this review. For the reconstitution of Adiantum neo1-LOV2 (*Ac neo1), we used the P905–P1087 construct containing the CBP tag at the N-terminus. For the comparison of the effect of Jα helix, we used the constructs encoding the same region of the LOV2-core and LOV2-Jα for Arabidopsis (At) phot1 and phot2 and Adiantum (Ac) neo1, where the GST tag at the N-terminus is digested with thrombin. (b) Amino acid sequences of the C-terminal side of LOV2-core and LOV2-Jα. This figure is modified from Ref 53.

Mentions: We have investigated light-induced structural changes of LOV domains by means of FTIR spectroscopy34,37–41,50–53. 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 samples51. We also describe the comprehensive FTIR analysis of LOV2 domains among Arabidopsis phot1, phot2, and Adiantum neo1 with and without Jα helix domain52,53 (Fig. 2).


Light-induced structural changes of the LOV2 domains in various phototropins revealed by FTIR spectroscopy
(a) Schematic illustration of the LOV2 constructs of phototropin presented in this review. For the reconstitution of Adiantum neo1-LOV2 (*Ac neo1), we used the P905–P1087 construct containing the CBP tag at the N-terminus. For the comparison of the effect of Jα helix, we used the constructs encoding the same region of the LOV2-core and LOV2-Jα for Arabidopsis (At) phot1 and phot2 and Adiantum (Ac) neo1, where the GST tag at the N-terminus is digested with thrombin. (b) Amino acid sequences of the C-terminal side of LOV2-core and LOV2-Jα. This figure is modified from Ref 53.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5036776&req=5

f2-7_89: (a) Schematic illustration of the LOV2 constructs of phototropin presented in this review. For the reconstitution of Adiantum neo1-LOV2 (*Ac neo1), we used the P905–P1087 construct containing the CBP tag at the N-terminus. For the comparison of the effect of Jα helix, we used the constructs encoding the same region of the LOV2-core and LOV2-Jα for Arabidopsis (At) phot1 and phot2 and Adiantum (Ac) neo1, where the GST tag at the N-terminus is digested with thrombin. (b) Amino acid sequences of the C-terminal side of LOV2-core and LOV2-Jα. This figure is modified from Ref 53.
Mentions: We have investigated light-induced structural changes of LOV domains by means of FTIR spectroscopy34,37–41,50–53. 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 samples51. We also describe the comprehensive FTIR analysis of LOV2 domains among Arabidopsis phot1, phot2, and Adiantum neo1 with and without Jα helix domain52,53 (Fig. 2).

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.