Periodic Reporting for period 1 - SmartSAST (Non-covalent photoresponsive tags for photoacoustic imaging: Giving voice to living matter)
Reporting period: 2020-04-01 to 2022-03-31
For Publication: " Long-Lived Charge-Transfer State from B–N Frustrated Lewis Pairs Enchained in Supramolecular Copolymers" (published Sept 2020)
Abstract: The field of supramolecular polymers is rapidly expanding; however, the exploitation of these systems as functional materials is still elusive. To become competitive, supramolecular polymers must display microstructural order and the emergence of new properties upon copolymerization. To tackle this, a greater understanding of the relationship between monomers’ design and polymer microstructure is required as well as a set of functional monomers that efficiently interact with one another to synergistically generate new properties upon copolymerization. Here, we present the first implementation of frustrated Lewis pairs into supramolecular copolymers. Two supramolecular copolymers based on π-conjugated O-bridged triphenylborane and two different triphenylamines display the formation of B–N pairs within the supramolecular chain. The remarkably long lifetime and the circularly polarized nature of the resulting photoluminescence emission highlight the possibility to obtain an intermolecular B–N charge transfer. These results are proposed to be the consequences of the enchainment of B–N frustrated Lewis pairs within 1D supramolecular aggregates. Although it is challenging to obtain a precise molecular picture of the copolymer microstructure, the formation of random blocklike copolymers could be deduced from a combination of optical spectroscopic techniques and theoretical simulation.
For publication: "A quantitative model for reversibly photoswitchable sensors" (in revision)
Abstract: Composed by a reversibly photoswitchable unit allosterically linked to a sensing module, reversibly photoswitchable sensors (rs-sensors) represent a new and attractive strategy to quantitatively read-out analyte concentrations. However their kinetic response to illumination is complex and
much attention is required from the design to the application steps. Here we exploit a generic kinetic model of rs-sensors, which enables us to point on key thermokinetic parameters such as dissociation constants and kinetic rates for exchange towards the analyte, and cross sections for
photoswitching. The application of the model allows to evaluate the robustness of the analyzed parameters and to introduce a methodology for their reliable use. Model and methodology have been experimentally tested on a newly reported calcium sensor based on a reversibly photoswitchable green fluorescent protein allosterically linked to a calcium-sensing module integrating calmoduline and a RS20 peptide.
Importance for the society: Importance in the field of chemistry, biology, microscopy and biophysics. The work will give a solid support to reliably exploit reversibly photoswitchable sensors
Objective within the smartSAST project: Although studied on a different protein-small molecule system, the publication details a theroetical and methodological section aimed to adress the issue of photo-ejection and photo-capture that will be used also to understand and employ the behavior of smartSAST sensors.
Abstract: The field of supramolecular polymers is rapidly expanding; however, the exploitation of these systems as functional materials is still elusive. To become competitive, supramolecular polymers must display microstructural order and the emergence of new properties upon copolymerization. To tackle this, a greater understanding of the relationship between monomers’ design and polymer microstructure is required as well as a set of functional monomers that efficiently interact with one another to synergistically generate new properties upon copolymerization. Here, we present the first implementation of frustrated Lewis pairs into supramolecular copolymers. Two supramolecular copolymers based on π-conjugated O-bridged triphenylborane and two different triphenylamines display the formation of B–N pairs within the supramolecular chain. The remarkably long lifetime and the circularly polarized nature of the resulting photoluminescence emission highlight the possibility to obtain an intermolecular B–N charge transfer. These results are proposed to be the consequences of the enchainment of B–N frustrated Lewis pairs within 1D supramolecular aggregates. Although it is challenging to obtain a precise molecular picture of the copolymer microstructure, the formation of random blocklike copolymers could be deduced from a combination of optical spectroscopic techniques and theoretical simulation.
For publication: "A quantitative model for reversibly photoswitchable sensors" (in revision)
Abstract: Composed by a reversibly photoswitchable unit allosterically linked to a sensing module, reversibly photoswitchable sensors (rs-sensors) represent a new and attractive strategy to quantitatively read-out analyte concentrations. However their kinetic response to illumination is complex and
much attention is required from the design to the application steps. Here we exploit a generic kinetic model of rs-sensors, which enables us to point on key thermokinetic parameters such as dissociation constants and kinetic rates for exchange towards the analyte, and cross sections for
photoswitching. The application of the model allows to evaluate the robustness of the analyzed parameters and to introduce a methodology for their reliable use. Model and methodology have been experimentally tested on a newly reported calcium sensor based on a reversibly photoswitchable green fluorescent protein allosterically linked to a calcium-sensing module integrating calmoduline and a RS20 peptide.
Importance for the society: Importance in the field of chemistry, biology, microscopy and biophysics. The work will give a solid support to reliably exploit reversibly photoswitchable sensors
Objective within the smartSAST project: Although studied on a different protein-small molecule system, the publication details a theroetical and methodological section aimed to adress the issue of photo-ejection and photo-capture that will be used also to understand and employ the behavior of smartSAST sensors.
For Publication: " Long-Lived Charge-Transfer State from B–N Frustrated Lewis Pairs Enchained in Supramolecular Copolymers"
Analysis of the data, drafting and reviewing of the paper.
For Publication: "A quantitative model for reversibly photoswitchable sensors" (in revision)
- Production and purification of the protein emplyed
- Photophysical and photochemical analysis of the behavior of the protein once submitted at difference calcium concentrations.
- We detailed the theoretical framework to understand the interconnected behavior of the protein's calcium complexation with the one of photoisomerization. The study displays how variation on applied light intensity may modify the working regime of the protein modifying its response and calcium complexation.
Analysis of the data, drafting and reviewing of the paper.
For Publication: "A quantitative model for reversibly photoswitchable sensors" (in revision)
- Production and purification of the protein emplyed
- Photophysical and photochemical analysis of the behavior of the protein once submitted at difference calcium concentrations.
- We detailed the theoretical framework to understand the interconnected behavior of the protein's calcium complexation with the one of photoisomerization. The study displays how variation on applied light intensity may modify the working regime of the protein modifying its response and calcium complexation.
For publication: "A quantitative model for reversibly photoswitchable sensors" (to be submitted soon)
Abstract: This manuscript proposes a theoretical frame and a methodology to ensure an educated characterization and use of the newlyintroduced class of rs-sensors, which provide an appealing alternative to state-of-the-art sensors.
We first introduce a generic kinetic model incorporating noncovalent binding of the analyte to the sensor whose reversible photoswitchable unit photochemical interconverts between two isomeric states. The exploitation of this model points on the significance of the threshold of light intensities at which photoswitching
and analyte exchange occur at the same rate. In this way, it allows to determine the optimal experimental conditions to exploit the time response of the sensor’s signal to illumination. By enabling the identification of regimes of light intensities and analyzed kinetic windows, the model further allows to reliably extract the
analyte concentration. Based on the model, we then propose a simple step-by-step experimental protocol, which provides the kinetic information to calibrate and exploit rs-sensors from robust mono- or bi-exponential fits after preliminarily analyzing the lightintensity dependence of the time response of the rs-sensor’s signal
at asymptotic analyte concentrations. Both kinetic model and characterization methodology have been applied on the newly reported calcium rs-sensor GCaMP6s-Q. In particular, we established that GCaMP6s-Q is compatible with illumination promoting essentially complete photo-conversions facilitating its preliminary calibration. Indeed the initial and final fluorescence signals of its time response to illumination depend only on the calcium concentration. We further showed that it favorably permits to avoid the introduction of any significant perturbation under calcium sensing.
We expect the kinetic model and the characterization procedure here proposed to be a fruitful support for the sensor developers by guiding them on the fundamental parameters to investigate for the future development of robust and versatile rs-sensors. Analogously, end-users will benefit from applying the characterization
procedure as it allows for reliable and quantitative analyte measurements
Abstract: This manuscript proposes a theoretical frame and a methodology to ensure an educated characterization and use of the newlyintroduced class of rs-sensors, which provide an appealing alternative to state-of-the-art sensors.
We first introduce a generic kinetic model incorporating noncovalent binding of the analyte to the sensor whose reversible photoswitchable unit photochemical interconverts between two isomeric states. The exploitation of this model points on the significance of the threshold of light intensities at which photoswitching
and analyte exchange occur at the same rate. In this way, it allows to determine the optimal experimental conditions to exploit the time response of the sensor’s signal to illumination. By enabling the identification of regimes of light intensities and analyzed kinetic windows, the model further allows to reliably extract the
analyte concentration. Based on the model, we then propose a simple step-by-step experimental protocol, which provides the kinetic information to calibrate and exploit rs-sensors from robust mono- or bi-exponential fits after preliminarily analyzing the lightintensity dependence of the time response of the rs-sensor’s signal
at asymptotic analyte concentrations. Both kinetic model and characterization methodology have been applied on the newly reported calcium rs-sensor GCaMP6s-Q. In particular, we established that GCaMP6s-Q is compatible with illumination promoting essentially complete photo-conversions facilitating its preliminary calibration. Indeed the initial and final fluorescence signals of its time response to illumination depend only on the calcium concentration. We further showed that it favorably permits to avoid the introduction of any significant perturbation under calcium sensing.
We expect the kinetic model and the characterization procedure here proposed to be a fruitful support for the sensor developers by guiding them on the fundamental parameters to investigate for the future development of robust and versatile rs-sensors. Analogously, end-users will benefit from applying the characterization
procedure as it allows for reliable and quantitative analyte measurements