Reversible Conformational Change Research Articles

Unveiling the Mechanism of Deprotonation and Proton Transfer of DNA Polymerase Catalysis via Single-Molecule Conductance.

DNA polymerases (Pols) play important roles in the transmission of genetic information. Although the function and (de)regulation of Pols are linked to many human diseases, the key mechanism of 3'-OH deprotonation and the PPi formation are not totally clear. In this work, a method is presented to detect the full catalytic cycle of human Pol (hPol β) in graphene-molecule-graphene single-molecule junctions. Real-time in situ monitoring successfully revealed the spatial and temporal properties of the open and closed conformation states of hPol β, distinguishing the reaction states in the Pols catalytic cycle and unveiling 3'-OH deprotonation and pyrophosphate (PPi) formation mechanism of hPol β. Proton inventory experiment demonstrated that the rate-limiting step of PPi formation is deprotonation, which occurs before a reverse conformational change. Additionally, by detecting the acidity (pKa), it is found that MgA-bound OH- acted as a general base and activated the nucleophile of 3'-OH, and that acidic residue D190 or D192 coordinated with MgB as a proton donor to PPi. This work provides useful insights into a fundamental chemical reaction that impacts genome synthesis efficiency and Pol fidelity, which the discovery of Pol-targeting drugs and design of artificial Pols for DNA synthetic applications are expected to accelerated.

Effect of Solvent Properties on the Critical Solution Temperature of Thermoresponsive Polymers.

The ability of thermoresponsive polymers to respond to temperature with a reversible conformational change makes them promising 'smart' materials for solutions in medical and biotechnological applications. In this work, two such polymers and structural isomers were studied: poly(N-isopropyl acrylamide) (PNiPAm) and poly(2-isopropyl-2-oxazoline) (PiPOx). We compare the critical solution temperatures (CST) of these polymers in D2O and H2O in the presence of Hofmeister series salts, as results obtained under these different solvent conditions are often compared. D2O has a higher dipole moment and electronegativity than H2O, which could significantly alter the CST transition. We used two complementary methods to measure the CST, dynamic light scattering (DLS) and differential scanning calorimetry (DSC) and found that the CST decreased significantly in D2O compared to H2O. In the presence of highly concentrated kosmotropes, the CST of both polymers decreased in both solvents. The influence of the kosmotropic anions was smaller than the water isotope effect at low ionic strengths but considerably higher at physiological ionic strengths. However, the Hofmeister anion effect was quantitatively different in H2O than in D2O, with the largest relative differences observed for Cl-, where the CSTs in D2O decreased more than in H2O measured by DLS but less by DSC. PiPOx was more sensitive than PNiPAm to the presence of chaotropes. It exhibited much higher transition enthalpies and multistep transitions, especially in aqueous solutions. Our results highlight that measurements of thermoresponsive polymer properties in D2O cannot be compared directly or quantitatively to application conditions or even measurements performed in H2O.

PH-Responsive, Wide Color Gamut Dynamic Color Display Enabled by PDMAEMA Brush-Based Fabry-Perot Resonant Cavity.

Dynamic color-changing materials have attracted broad interest due to their widespread applications in visual sensing, dynamic color display, anticounterfeiting, and image encryption/decryption. In this work, we demonstrate a novel pH-responsive dynamic color-changing material based on a metal-insulator-metal (MIM) Fabry-Perot (FP) cavity with a pH-responsive poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA) brush layer as the responsive insulating layer. The pH-responsive PDMAEMA brush undergoes protonation at a low pH value (pH < 6), which induces different swelling degrees in response to pH and thus refractive index and thickness change of the insulator layer of the MIM FP cavity. This leads to significant optical property changes in transmission and a distinguishable color change spanning the whole visible region by adjusting the pH value of the external environment. Due to the reversible conformational change of the PDMAEMA and the formation of covalent bonds between the PDMAEMA molecular chain and the Ag substrate, the MIM FP cavity exhibits stable performance and good reproducibility. This pH-responsive MIM FP cavity establishes a new way to modulate transmission color in the full visible region and exhibits a broad prospect of applications in dynamic color display, real-time environment monitoring, and information encryption and decryption.

Conformational photo-trapping in NaV1.5: Inferring local motions at the “inactivation gate”

Rapid and effectual inactivation in voltage-gated sodium channels is required for canonical action-potential firing. This “fast” inactivation arises from swift and reversible protein conformational changes that utilize transmembrane segments and the cytoplasmic linker between channel domains III and IV. Until recently, fast inactivation had been accepted to rely on a “ball-and-chain” mechanism whereby a hydrophobic triplet of DIII-IV amino acids (IFM) impairs conductance by binding to a site in central pore of the channel made available by channel opening. New structures of sodium channels have upended this model. Specifically, cryo-electron microscopic structures of eukaryotic sodium channels depict a peripheral binding site for the IFM motif, outside of the pore, opening the possibility of a yet unidentified allosteric mechanism of fast-inactivation gating. We set out to study fast inactivation by photo-trapping human sodium channels in various functional states under voltage control. This was achieved by genetically encoding the crosslinking unnatural amino acid benzophenone phenylalanine at various sites within the DIII-IV linker in the cardiac sodium channel NaV1.5. These data show dynamic state- and positional-dependent trapping of the transient conformations associated with fast inactivation, each yielding different phenotypes and rates of trapping. These data reveal distinct conformational changes that underlie fast inactivation and point to a dynamic environment around the IFM locus.

Reprogrammable Multi‐Responsiveness of Regenerated Silk for Versatile Soft Actuators

AbstractDeveloping stimulus‐responsive materials (SRMs) with reversible, large, and reprogrammable multi‐responsiveness is crucial for soft actuators and intelligent devices, which remains challenging. In this study, regenerated silk fibroin with hierarchical structure is utilized to specially design a reprogrammable multi‐stimuli‐responsive protein film and versatile soft actuators. The freestanding fibroin films exhibit notable thermal contraction (−1383 ppm K−1) and humidity‐responsiveness that can be repeatedly regulated by easy post‐treatment with Ca2+ as desired. The actuating and regulating mechanisms involve reversible conformational change that is magnified into macroscopic deformations by hierarchical structure, and the roles of material structure and ambient conditions in determining actuating performances are analyzed based on thermodynamics. Driven by humidity gradient, the fibroin film demonstrates spontaneous flipping locomotion, self‐oscillation with tunable frequencies, bio‐butterfly wing flapping, and transformation from 2D to 3D structure. Moreover, reprogrammable deformations at specific regions are achieved in multi‐stimuli‐driven PET/fibroin film actuators owing to the straightforward Ca2+‐content‐based tunability of the responsiveness. The fibroin‐based actuators can be used as artificial muscles to drive the high‐frequency wing‐flapping of a bio‐dragonfly and soft gripper to grasp, lift, and transfer objects. The simple yet effective strategy presented herein provides valuable inspiration for designing advanced SRMs and soft actuators with reprogrammable multi‐responsiveness.

Visible light-mediated supramolecular framework for tunable CO2 adsorption

In the pursuit of creating azobenzene-functionalized photo-responsive supramolecular frameworks (PSMFs) activated by the visible spectrum, an endeavor traditionally relied on ultraviolet activation, we report the successful fabrication of a visible light-mediated metal–organic cage-based PSMF, designated as NUT-104. This achievement was realized by the hierarchical self-assembly process of Zr-metal–organic cages featuring ortho-fluoroazobenzene functional terephthalic acid and tri-nuclear Zr-clusters. NUT-104 displays permanent porosity coupled with reversible responsiveness to visible light. This remarkable behavior emerges as a consequence of the σ-electron-withdrawing effect induced by the presence of fluorine substituents. Visible light-regulated CO2 adsorption capacity can be achieved and the change value can reach 61 %, while the change for CH4 and N2 is only 7 % and 3 %, respectively. In response to irradiation at wavelengths of 520 and 420 nm, respectively, the azobenzene moieties within the NUT-104 structure exhibit a remarkable capacity for visible light-controlled reversible conformational changes. Grand Canonical Monte Carlo simulations unveiled that the dynamic reorganization of the NUT-104 proficiently governs the accessibility of CO2 adsorption sites positioned within the interstitial regions of proximate cages. The advent of NUT-104, underpinned by its visible light responsiveness and versatile tunability, paves the way for avenues of exploration in controlled gas separation and storage applications.

Unlocking giant negative thermal expansion through reversible conformational changes in regenerated silk fibroin film

Developing thermally contractive polymeric materials with controlled conformational changes that enable more efficient actuation mechanisms has long been an ambitious goal. However, the utilization of naturally derived assemblies with biodegradable and reversible capabilities has, thus far, been hindered by the challenges associated with organizing the hierarchical nanostructures. Herein, hierarchically structured building blocks in regenerated fibroin were utilized to construct a thermal-responsive protein film through a friction-induced assemble strategy. By preserving the protofibrils within the regenerated silk film, we enable the efficient storage and transfer of elastic energy to external loads. The freestanding fibroin film, with pre-extended molecular chains, thus produced exhibited an exceedingly high negative thermal expansion (−1220 ppm K−1) and work capacity (∼203 J kg−1). The comprehensive analysis involving synchrotron infrared spectroscopy, in-situ Raman spectroscopy, and molecular dynamics simulations has confirmed that the actuating mechanism entails a reversible conformational change initiated by H-bonds, which is then amplified into macroscopic deformations by the hierarchical structure. This newly discovered, low-energy-driven mechanism paves the way for the creation of flexible protein assemblies with high-performance actuation capabilities.

Thermoresponsive membrane based on UCST-type organoboron polymer for smart gating and self-cleaning

Thermoresponsive polymers can endow membranes with tunable separation and self-cleaning performance. However, up to now, there have been only very limited choices of thermoresponsive polymers for the fabrication of smart membranes. Herein, thermoresponsive membranes based on UCST-type organoboron polymer (Poly(CPBA)) were fabricated for the first time. Poly(CPBA) synthesized via a nucleophilic ring-opening reaction with Poly(GMA) and 3-carboxyphenylboronic acid has shown typical thermoresponsive behavior in water and water/ethanol mixture. Under vacuum conditions, Poly(CPBA) formed the cross-linked boroxine network on the membrane surface via a dehydration-induced condensation reaction. The reversible conformational change of Poly(CPBA) near UCST acted as a smart “gating” to regulate both pore size and surface properties of the membrane. When the temperature exceeded UCST, the fully extended polymer chains provided a strong washing force to remove contaminants, resulting in effective self-cleaning performance (FRR up to 99.2 %). In particular, due to the presence of the boroxine network, the polymer chains were firmly “stuck” on the membrane surface, providing stable performance and superior repeatability of the composite membrane.

Terahertz characterization of combined pressure-shear shock loaded aromatic polyurea

Realistic shock loading scenarios imply combined, multiaxial stress states, where the mechanically induced energy competes for failing the material intrinsically due to cohesive failures. Under shock loading, failure encompasses various modes, including spallation, shear bands, microcracking, morphological alteration, phase transformation, and plastic deformation. However, elucidating failure in high strain loading conditions challenges the state-of-the-art characterization due to the disparity interfacing different loading and measurement techniques. This research used a novel spectro-mechanical characterization setup, integrating bulk terahertz spectroscopy synchronized in real-time with mixed-mode laser-induced shock waves. The latter was used to submit polyurea elastomers to concurrent pressure and shear waves by saddling a thin polymer sheet on the hypotenuse of a glass prism. Hence, the laser-induced pressure shock waves underwent simultaneous mode conversion at the glass-polyurea interface at the hypotenuse of the glass prism, effectively submitting the sample to tunable amplitude pressure-shear shock waves. The terahertz spectroscopy, operating in the right-angle reflection configuration, was modified to probe predetermined spots on the polyurea samples at different stress states (adjusted by changing the illumination energy) in a three-step sequence: before, during, and after shock loading. The shock-loaded polyurea samples were also characterized using the optical and scanning electron microscopes, revealing compression-shear and tensile-shear failure modes, including surface depression, nucleation of micro-voids and microcracks, and formation of small planes. The induced stress state based on the illumination energy was quantified using an accompanying finite element simulation, resulting in completing the quasi-isentropic behavior of polyurea elastomers, which was found to be in good agreement with previous experimental data from the plate impact experiments and the predictions based on the Lennard-Jones potential. Time domain analysis of terahertz data, using temporal shifting, hysteresis, and dynamic time warping based on the sum of the Euclidean distances between signal pairs, affirmed the microscopy observations while shedding insights on the elastic and plastic deformations and the reversible conformational changes. Irreversible changes to the macromolecule were also detected using the terahertz data spectral analysis. The current results establish a new experimental framework for fundamentally characterizing polymers and their composites at ultrahigh strain rate loading conditions.

Molecular machine-based nanomaterials: Recent progress and prospects

Molecular machines are chemical molecules (man-made or natural) which display triggered reversible conformational changes by external stimuli like pH and temperature, followed by observable changes. This property of controllable and coherent motion at molecular level can be observed through the synthesis of macroscopic materials from self-assembly of molecular machines. Due to these properties, molecular machine-based nanomaterials display a wide range of unique characteristics and various useful applications in biomedicine, energy, environmental engineering, etc. This paper reviews recent research articles on some potential applications including biosensing, therapeutics (with a focus on drug delivery system) and catalysis based on MMs and their composites to assess the current practicality of the technology. Although most materials are not ready for real-life applications yet and there is a lack of deep understanding of the efficacy, these nanomaterials display a wide range of very useful and unique characteristics and present a brand-new direction for tackling current challenges.

Adsorption and reversible conformational change of a thiophene based molecule on Au(111)

We present a low temperature scanning tunneling microscope investigation of a prochiral thiophene-based molecule that self-assembles forming islands with different domains on the Au(111) surface. In the domains, two different conformations of the single molecule are observed, depending on a slight rotation of two adjacent bromothiophene groups. Using voltage pulses from the tip, single molecules can be switched between the two conformations. The electronic states have been measured with scanning tunneling spectroscopy, showing that the electronic resonances are mainly localized at the same positions in both conformations. Density-functional theory calculations support the experimental results. Furthermore, we observe that on Ag(111), only one configuration is present and therefore the switching effect is suppressed.

Time-resolved investigation of nanometric cell membrane patches with a mid-infrared laser microscope

The proton pump Bacteriorhodopsin (BR) undergoes repeated photocycles including reversible conformational changes upon visible light illumination. Exploiting the sensitivity of infrared (IR) spectra to the conformation, we have determined the reaction kinetic parameters of the conductive intermediate M for the wild-type protein and for its slow mutant D96N during its photocycle. Time-resolved IR micro-spectroscopy using an in-house developed confocal laser microscope operating in the mid-IR is employed to record absorption changes of 10−4 at wavelengths λ1 = 6.08 μm and λ2 = 6.35 μm, assigned to backbone and retinal structural modifications, respectively. Protein samples were embedded in dried lipid bilayers deposited on ultraflat gold supports to enhance the surface field. The signals were analyzed according to a simplified photocycle model with only two dominant states: the dark-adapted state BR* and the intermediate M. We obtained the excitation and relaxation times of the intermediate M from exponential fits to the absorption change time traces. Our results constitute a first step towards future plasmonic-assisted nanoscale time-resolved mid-IR spectrometers for the characterization of bioelectronic and light-harvesting nanodevices based on BR.

Uncovering allostery and regulation in SORCIN through molecular dynamics simulations

Soluble resistance-related calcium-binding protein or Sorcin is an allosteric, calcium-binding Penta-EF hand (PEF) family protein implicated in multi-drug resistant cancers. Sorcin is known to bind chemotherapeutic molecules such as Doxorubicin. This study uses in-silico molecular dynamics simulations to explore the dynamics and allosteric behavior of Sorcin in the context of Ca2+ uptake and Doxorubicin binding. The results show that Ca2+ binding induces large, but reversible conformational changes in the Sorcin structure which manifest as rigid body reorientations that preserve the local secondary structure. A reciprocal allosteric handshake centered around the EF5 hand is found to be key in Sorcin dimer formation and stabilization. Binding of Doxorubicin results in rearrangement of allosteric communities which disrupts long-range allosteric information transfer from the N-terminal domain to the middle lobe. However, this binding does not result in secondary structure destabilization. Sorcin does not appear to have a distinct Ca2+ activated mode of Doxorubicin binding. Communicated by Ramaswamy H. Sarma

An ensemble of interconverting conformations of the elemental paused transcription complex creates regulatory options.

Transcriptional pausing underpins the regulation of cellular RNA synthesis, but its mechanism remains incompletely understood. Sequence-specific interactions of DNA and RNA with the dynamic, multidomain RNA polymerase (RNAP) trigger reversible conformational changes at pause sites that temporarily interrupt the nucleotide addition cycle. These interactions initially rearrange the elongation complex (EC) into an elemental paused EC (ePEC). ePECs can form longer-lived PECs by further rearrangements or interactions of diffusible regulators. For both bacterial and mammalian RNAPs, a half-translocated state in which the next DNA template base fails to load into the active site appears central to the ePEC. Some RNAPs also swivel interconnected modules that may stabilize the ePEC. However, it is unclear whether swiveling and half-translocation are requisite features of a single ePEC state or if multiple ePEC states exist. Here, we use cryo-electron microscopy (cryo-EM) analysis of ePECs with different RNA-DNA sequences combined with biochemical probes of ePEC structure to define an interconverting ensemble of ePEC states. ePECs occupy either pre- or half-translocated states but do not always swivel, indicating that difficulty in forming the posttranslocated state at certain RNA-DNA sequences may be the essence of the ePEC. The existence of multiple ePEC conformations has broad implications for transcriptional regulation.

De novo sequencing and construction of a unique antibody for the recognition of alternative conformations of cytochrome c in cells

Cytochrome c (cyt c) can undergo reversible conformational changes under biologically relevant conditions. Revealing these alternative cyt c conformers at the cell and tissue level is challenging. A monoclonal antibody (mAb) identifying a key conformational change in cyt c was previously reported, but the hybridoma was rendered nonviable. To resurrect the mAb in a recombinant form, the amino-acid sequences of the heavy and light chains were determined by peptide mapping-mass spectrometry-bioinformatic analysis and used to construct plasmids encoding the full-length chains. The recombinant mAb (R1D3) was shown to perform similarly to the original mAb in antigen-binding assays. The mAb bound to a variety of oxidatively modified cyt c species (e.g., nitrated at Tyr74 or oxidized at Met80), which lose the sixth heme ligation (Fe-Met80); it did not bind to several cyt c phospho- and acetyl-mimetics. Peptide competition assays together with molecular dynamic studies support that R1D3 binds a neoepitope within the loop 40-57. R1D3 was employed to identify alternative conformations of cyt c in cells under oxidant- or senescence-induced challenge as confirmed by immunocytochemistry and immunoaffinity studies. Alternative conformers translocated to the nuclei without causing apoptosis, an observation that was further confirmed after pinocytic loading of oxidatively modified cyt c to B16-F1 cells. Thus, alternative cyt c conformers, known to gain peroxidatic function, may represent redox messengers at the cell nuclei. The availability and properties of R1D3 open avenues of interrogation regarding the presence and biological functions of alternative conformations of cyt c in mammalian cells and tissues.

Polymer elastomer near plastic-to-rubber critical transition produces enhanced elastocaloric effects

Solid-state cooling technology based on elastocaloric effects has shown potential advantages over traditional vapor-compression cooling systems owing to its high energy efficiency and zero greenhouse gas emissions. However, a lack of strategies for improving the elastocaloric activity of polymers remains the main obstacle to their commercialization since the elastocaloric effect was first investigated by J.P. Joule in natural rubber. Herein, we demonstrate that polymer elastomers with molecular chains near molecular-structure-induced plastic-rubber critical transitions exhibit superior elastocaloric activity through reversible conformational changes. As the content of ethyl side groups in the soft block of polystyrene- b -poly(ethylene- co -butylene)- b -polystyrene decreases from 33 to 8 mol %, the adiabatic temperature change increases from −4.3 to −16.3 K, and the isothermal entropy change increases from 38.5 to 173.3 J kg −1 K −1 , which thus shows the highest polymer elastocaloric activity near room temperature. This strategy could promote the commercialization of elastocaloric polymers in solid-state cooling applications. • Polymers near plastic-rubber critical transition show reversible conformations • Over a 3-fold increment in the elastocaloric activity close to room temperature • The lowest limit value of cooling temperature decreased from 275 to 262 K Solid-state cooling technologies are expected to be a net-zero potential alternative to vapor-compression cooling systems. Here, Shixian Zhang et al. demonstrate a strategy to enhance the cooling performance of solid polymers. This strategy could be widely applied in polymer systems to reach polymers’ solid-state cooling.

Photoactivation of a Mechanosensitive Channel.

Optogenetics in the conventional sense, i.e. the use of engineered proteins that gain their light sensitivity from naturally abundant chromophores, represents an exciting means to trigger and control biological activity by light. As an alternate approach, photopharmacology controls biological activity with the help of synthetic photoswitches. Here, we used an azobenzene-derived lipid analogue to optically activate the transmembrane mechanosensitive channel MscL which responds to changes in the lateral pressure of the lipid bilayer. In this work, MscL has been reconstituted in nanodiscs, which provide a native-like environment to the protein and a physical constraint to membrane expansion. We characterized this photomechanical system by FTIR spectroscopy and assigned the vibrational bands of the light-induced FTIR difference spectra of the trans and cis states of the azobenzene photolipid by DFT calculations. Differences in the amide I range indicated reversible conformational changes in MscL as a direct consequence of light switching. With the mediation of nanodiscs, we inserted the transmembrane protein in a free standing photoswitchable lipid bilayer, where electrophysiological recordings confirmed that the ion channel could be set to one of its sub-conducting states upon light illumination. In conclusion, a novel approach is presented to photoactivate and control cellular processes as complex and intricate as gravitropism and turgor sensing in plants, contractility of the heart, as well as sensing pain, hearing, and touch in animals.

Phenanthrene-Extended Phenazine Dication: An Electrochromic Conformational Switch Presenting Dual Reactivity.

The synthesis and isolation of one of the few examples of a π-extended diamagnetic phenazine dication have been achieved by oxidizing a phenanthrene-based dihydrophenazine precursor. The resulting dication was isolated and fully characterized, highlighting an aromatic distorted structure, generated by the conformational change upon the oxidation of the dihydrophenazine precursor, which is also correlated with a marked electrochromic change in the UV–vis spectrum. The aromaticity of the dication has also been investigated theoretically, proving that the species is aromatic based on all major criteria (structural, magnetic, and energetic). Moreover, the material presents an intriguing dual reactivity, resulting in ring contraction to a π-extended triarylimidazolinium and reduction to the dihydrophenazine precursor, depending on the nature of the nucleophile involved. This result helps shed light on the yet largely unexplored reactivity and properties of extended dicationic polycyclic aromatic hydrocarbons (PAHs). In particular, the fact that the molecule can undergo a reversible change in conformation upon oxidation and reduction opens potential applications for this class of derivatives as molecular switches and actuators.

Molecular Switches-Tools for Imparting Control in Drug Delivery Systems.

Cancer is a globally prevalent cause of premature mortality. Of growing interest is the development of novel anticancer therapies and the optimisation of associated risks. Major issues presently facing conventional anticancer therapies include systemic toxicity, poor solubility, membrane permeability, and multidrug resistance Nanocarriers have been employed to address these issues. Nanocarriers encapsulate anticancer drugs, enabling them to bypass biological barriers and minimise their adverse side effects. These drug delivery systems offer extensive benefits as they can be modified to gravitate towards specific environmental conditions. To further enhance the safety and efficacy of these drug carriers, modern developments have included incorporating a molecular switching mechanism into their structure. These molecular switches are responsive to endogenous and exogenous stimuli and may undergo reversible and repeatable conformational changes when activated. The incorporation of molecular switches can, therefore, impart stimuli-responsive drug-release control on a DDS. These stimuli can then be manipulated to offer precise dosage control over the drug release at a specific target site. This review discusses recent developments in the design of DDSs incorporating light and pH-responsive molecular switches as drug release controllers.

A synthetic switch based on orange carotenoid protein to control blue-green light responses in chloroplasts.

Synthetic biology approaches to engineer light-responsive systems are widely used, but their applications in plants are still limited due to the interference with endogenous photoreceptors and the intrinsic requirement of light for photosynthesis. Cyanobacteria possess a family of soluble carotenoid-associated proteins named orange carotenoid proteins (OCPs) that, when activated by blue–green light, undergo a reversible conformational change that enables the photoprotection mechanism that occurs on the phycobilisome. Exploiting this system, we developed a chloroplast-localized synthetic photoswitch based on a protein complementation assay where two nanoluciferase fragments were fused to separate polypeptides corresponding to the OCP2 domains. Since Arabidopsis (Arabidopsis thaliana) does not possess the prosthetic group needed for the assembly of the OCP2 complex, we first implemented the carotenoid biosynthetic pathway with a bacterial β-carotene ketolase enzyme (crtW) to generate keto-carotenoid-producing plants. The photoswitch was tested and characterized in Arabidopsis protoplasts and stably transformed plants with experiments aimed to uncover its regulation by a range of light intensities, wavelengths, and its conversion dynamics. Finally, we applied the OCP-based photoswitch to control transcriptional responses in chloroplasts in response to green light illumination by fusing the two OCP fragments with the plastidial SIGMA FACTOR 2 and bacteriophage T4 anti-sigma factor AsiA. This pioneering study establishes the basis for future implementation of plastid optogenetics to regulate organelle responses upon exposure to specific light spectra.