Photosensitivity of Kinase Activation by Blue Light Involves the Lifetime of a Cysteinyl-Flavin Adduct Intermediate, S390, in the Photoreaction Cycle of the LOV2 Domain in Phototropin, a Plant Blue Light Receptor

Okajima, Koji; Kashojiya, Sachiko; Tokutomi, Satoru

doi:10.1074/jbc.m112.406512

Cited by 44 publications

(55 citation statements)

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“…Upon return to the dark, the adduct state spontaneously decays to an oxidized flavin (LOV 450 ) on a timescale of seconds to days (17,18). Currently the biological role of the wide range in photocycle lifetimes is unknown; however, several studies have suggested that the range facilitates adaptation to changing levels of light intensity (17,19,20). For these reasons, chemical tuning of the LOV photocycle lifetime through understanding of the adduct decay mechanism has been attempted in several systems (2,17,18,(21)(22)(23)(24).…”

Section: Light-oxygen-voltage (Lov)mentioning

confidence: 99%

A Native Threonine Coordinates Ordered Water to Tune Light-Oxygen-Voltage (LOV) Domain Photocycle Kinetics and Osmotic Stress Signaling in Trichoderma reesei ENVOY

Lokhandwala

Vega

Hopkins

et al. 2016

Journal of Biological Chemistry

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Light-oxygen-voltage (LOV)3 domain-containing photoreceptors are widely distributed in nature, where they couple blue light absorption to regulation of a diverse array of signal transduction pathways (1). In general, LOV proteins can be divided into two subclasses: 1) short LOV proteins (sLOV) that exist as the isolated LOV domain with short ancillary N-or C-terminal caps (2-4) and 2) modular LOV proteins that couple blue light activation to allosteric regulation of effector domains (1). Although the modular LOV proteins are present in plants, bacteria, and all fungi, the sLOV variety are predominantly found in bacteria and only some fungi (4). Currently, LOV proteins have received widespread attention because of their important roles in regulation of circadian function (3, 5, 6), growth and development (7,8), stress responses (9 -11), and adaptation to and regulation of pathogenicity (12). In addition, their wide ranging utility has led to the development of optogenetic tools that harness their modular design (13,14). Despite substantial research, key questions involving LOV photocycles remain.LOV domain chemistry is characterized by blue light-induced formation of a covalent adduct between a bound flavin cofactor (FMN, FAD, or riboflavin) and a conserved Cys residue in a GXNCRFLQ motif. Concomitant with adduct formation is protonation of the N5 position of the isoalloxazine ring. Current models indicate that signal transduction is coupled to N5 protonation via allosteric regulation of N-or C-terminal effector elements remote from the flavin active site (3,15,16). The covalent adduct is defined by a broad UV-visible absorption band centered at ϳ390 nm (LOV 390 ). Upon return to the dark, the adduct state spontaneously decays to an oxidized flavin (LOV 450 ) on a timescale of seconds to days (17,18). Currently the biological role of the wide range in photocycle lifetimes is unknown; however, several studies have suggested that the range facilitates adaptation to changing levels of light intensity (17,19,20). For these reasons, chemical tuning of the LOV photocycle lifetime through understanding of the adduct decay mechanism has been attempted in several systems (2,17,18,(21)(22)(23)(24).Several lines of reasoning have led to a general mechanism of adduct decay. First, solvent isotope effect experiments indicate that a single proton abstraction event is rate-limiting (17, 18). Second, adduct decay can be catalyzed by the presence of small molecule bases such as imidazole (25). Third, residue substitutions at regions that regulate solvent access to the flavin active site have a substantial effect on LOV photocycle lifetimes (18,26). Combined, these experiments implicate N5 deprotonation as the rate-determining step in adduct decay. Consistent with such a model, mutation of residues that regulate accessibility of small molecules to the N5 position or that tune hydrogen bonding characteristics affect kinetics of LOV proteins (17,18,21,22,24,26,27). Importantly, the natural base responsible for N5 deprotonation remai...

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Section: Light-oxygen-voltage (Lov)mentioning

confidence: 99%

A Native Threonine Coordinates Ordered Water to Tune Light-Oxygen-Voltage (LOV) Domain Photocycle Kinetics and Osmotic Stress Signaling in Trichoderma reesei ENVOY

Lokhandwala

Vega

Hopkins

et al. 2016

Journal of Biological Chemistry

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“…Within seconds to minutes, the S390 reverts to D450, thermally depending on the types of LOV (14), and forms a characteristic photocycle with LOV. S390 is a signaling state capable of activating the kinase (15). Of the two LOV domains, LOV2 has been shown to play a major role in kinase activation by BL through both autophosphorylation (16,17) and substrate phosphorylation in vitro (18).…”

mentioning

confidence: 99%

“…One of the main reasons for this problem is the difficulty in preparing phot samples consisting of both LOV and KD with a high enough purity and in large enough quantities for biophysical analyses. To overcome this difficulty, we previously established a preparation system for pure LOV2-KD of At phot1 and phot2 (15,26). The LOV2-KDs showed BL-regulated kinase activity on the N-terminal fragment of At phot1 (AtP1Nt), including the autophosphorylation sites around the LOV1 region that was created as an artificial substrate for the phot kinase (15,26).…”

mentioning

confidence: 99%

“…To overcome this difficulty, we previously established a preparation system for pure LOV2-KD of At phot1 and phot2 (15,26). The LOV2-KDs showed BL-regulated kinase activity on the N-terminal fragment of At phot1 (AtP1Nt), including the autophosphorylation sites around the LOV1 region that was created as an artificial substrate for the phot kinase (15,26). Because the LOV2-KDs have BL-regulated kinase activity, structural study of these molecules provides useful information regarding the molecular basis for the BL-dependent regulation of the kinase.…”

mentioning

confidence: 99%

“…Samples were run on SDS-PAGE. After Coomassie Brilliant Blue (CBB) staining, the gel was dried, and phosphorylated bands were visualized with an imaging plate (Fuji film) and a scanner (STORM, GE Healthcare) (15). The signal intensities were quantified by ImageJ software (rsbweb.…”

mentioning

confidence: 99%

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Light-induced Conformational Changes of LOV1 (Light Oxygen Voltage-sensing Domain 1) and LOV2 Relative to the Kinase Domain and Regulation of Kinase Activity in Chlamydomonas Phototropin

Okajima

Aihara

Takayama

et al. 2014

Journal of Biological Chemistry

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Background:The plant photoreceptor "phototropin" is a light-regulated kinase containing two photosensory domains named LOV. Results: Light-induced conformational change related to the kinase activation was detected in full-length phototropin of Chlamydomonas. Conclusion: LOV1 may interact with LOV2 and modify the photosensitivity of the kinase regulation by LOV2. Significance: Configuration of LOV1, LOV2, and kinase domain in a phot molecule is first demonstrated.

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The linker between LOV2‐Jα and STK plays an essential role in the kinase activation by blue light in Arabidopsis phototropin1, a plant blue light receptor

et al. 2015

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Edited by Richard CogdellPhototropin (phot), a blue light receptor in plants, is composed of several domains: LOV1, LOV2, and a serine/threonine kinase (STK). LOV2 is the main regulator of light activation of STK. However, the detailed mechanism remains unclear. In this report, we focused on the linker region between LOV2 and STK excluding the Ja-helix. Spectroscopy and a kinase assay for the substituents in the linker region of Arabidopsis phot1 LOV2-STK indicated that the linker is involved in the activation of STK. A putative module in the middle of the linker would be critical for intramolecular signaling and/or regulation of STK.

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Photosensitivity of Kinase Activation by Blue Light Involves the Lifetime of a Cysteinyl-Flavin Adduct Intermediate, S390, in the Photoreaction Cycle of the LOV2 Domain in Phototropin, a Plant Blue Light Receptor

Abstract: Background:The plant photoreceptor "phototropin" is a light-regulated kinase containing the LOV photosensory domains.

Cited by 44 publications

References 44 publications

A Native Threonine Coordinates Ordered Water to Tune Light-Oxygen-Voltage (LOV) Domain Photocycle Kinetics and Osmotic Stress Signaling in Trichoderma reesei ENVOY

A Native Threonine Coordinates Ordered Water to Tune Light-Oxygen-Voltage (LOV) Domain Photocycle Kinetics and Osmotic Stress Signaling in Trichoderma reesei ENVOY

Light-induced Conformational Changes of LOV1 (Light Oxygen Voltage-sensing Domain 1) and LOV2 Relative to the Kinase Domain and Regulation of Kinase Activity in Chlamydomonas Phototropin

The linker between LOV2‐Jα and STK plays an essential role in the kinase activation by blue light in Arabidopsis phototropin1, a plant blue light receptor

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