light-driven-molecular-motors-on-surfaces-for-single-molecular-imaging

Light-driven Molecular Motors on Surfaces for Single Molecular Imaging

The design and synthesis of a synthetic system that aims for the direct visualization of a synthetic rotary motor at the single molecule level on surfaces ...

imaging-guided-bioreactor-for-generating-bioengineered-airway-tissue

Imaging-Guided Bioreactor for Generating Bioengineered Airway Tissue

Repeated injury to airway tissue can impair lung function and cause chronic lung disease, such as chronic obstructive pulmonary disease. Advances in ...

University of Groningen

Stevens Institute of Technology

A series of first-generation light-driven molecular motors with rigid substituents of varying length was synthesized to act as "molecular stirrers". Their rotary motion was studied by (1)H NMR and UV-vis absorption spectroscopy in a variety of solvents with different polarity and viscosity. Quantitative analyses of kinetic and thermodynamic parameters show that the rotary speed is affected by the rigidity of the substituents and the length of the rigid substituents and that the differences in speed are governed by entropy effects. Most pronounced is the effect of solvent viscosity on the rotary motion when long, rigid substituents are present. The α values obtained by the free volume model, supported by DFT calculations, demonstrate that during the rotary process of the motor, as the rigid substituent becomes longer, an increased rearranging volume is needed, which leads to enhanced solvent displacement and retardation of the motor. 

Molecular stirrers in action.

The facile assembly of higher-order nanoarchitectures from simple building blocks is demonstrated by the loading of vesicles into soft amphiphilic nanotubes using osmosis. The nanotubes are constructed from rigid interdigitated bilayers which are capped with vesicles comprising phospholipid-based flexible bilayers. When a hyperosmotic gradient is applied to these vesicle-capped nanotubes, the closed system loses water and the more flexible vesicle bilayer is pulled inwards. This leads to inclusion of vesicles inside the nanotubes without affecting the tube structure, showing controlled reorganization of the self-assembled multicomponent system upon a simple osmotic stimulus. 

Loading of Vesicles into Soft Amphiphilic Nanotubes using Osmosis.

The novel concept of amphiphilic molecular motors that self-assemble into responsive supramolecular nanotubes in water is presented. The dynamic function of the molecular motor units inside the supramolecular assemblies was studied using UV-vis absorption spectroscopy and cryo-transmission electron microscopy (cryo-TEM) microscopy. Reorganization between distinct, well-defined nanotubes and vesicles can be reversibly induced by light, going through the rotation cycle of the motor, i.e. driven by alternate photochemical and thermal isomerization steps in the system. This is the first example in which a molecular rotary motor shows self-assembly in an aqueous medium with full retention of its functionality, paving the way to increasingly complex, highly dynamic artificial nanosystems in water. 

Amphiphilic Molecular Motors for Responsive Aggregation in Water.

An intramolecular cargo transport device, based on a light-driven molecular motor, is presented. Detailed UV-vis and (1)H NMR studies demonstrate that the gripper attached to the upper half of the molecular motor is able to pick up an acetyl group from one side of the lower stator and, after a photochemical Z-E isomerization, drop it on the opposite side.

Intramolecular transport of small-molecule cargo in a nanoscale device operated by light.

Soft amphiphilic nanotubes are capped with vesicles comprised of either overall neutral, zwitterionic phospholipids, or those that carry a net charge. The phase transition temperature of the zwitterionic phospholipids plays a crucial role in the phase separation that leads to the end-capped nanotubes. The cationic vesicle caps can be loaded into the nanotubes via osmosis whereas the anionic vesicle caps are stable under hyper-osmotic conditions. Furthermore, no additional salt needs to be added for the cationic vesicle caps to induce the loading of the vesicles into the nanotubes due to the presence of counterions.

End-capping of amphiphilic nanotubes with phospholipid vesicles: impact of the phospholipid on the cap formation and vesicle loading under osmotic conditions.

Photochemically driven molecular motors convert the energy of incident radiation to intramolecular rotational motion. The motor molecules considered here execute four step unidirectional rotational motion. This comprises a pair of successive light induced isomerizations to a metastable state followed by thermal helix inversions. The internal rotation of a large molecular unit required in these steps is expected to be sensitive to both the viscosity of the medium and the volume of the rotating unit. In this work, we describe a study of motor motion in both ground and excited states as a function of the size of the rotating units. The excited state decay is ultrafast, highly non-single exponential, and is best described by a sum of three exponential relaxation components. The average excited state decay time observed for a series of motors with substituents of increasing volume was determined. While substitution does affect the lifetime, the size of the substituent has only a minor effect. The solvent polarity dependence is also slight, but there is a significant solvent viscosity effect. Increasing the viscosity has no effect on the fastest of the three decay components, but it does lengthen the two slower decay times, consistent with them being associated with motion along an intramolecular coordinate displacing a large solvent volume. However, these slower relaxation times are again not a function of the size of the substituent. We conclude that excited state decay arises from motion along a coordinate which does not necessarily require complete rotation of the substituents through the solvent, but is instead more localized in the core structure of the motor. The decay of the metastable state to the ground state through a helix inversion occurs 14 orders of magnitude more slowly than the excited state decay, and was measured as a function of substituent size, solvent viscosity and temperature. In this case neither substituent size nor solvent viscosity influences the rate, which is entirely determined by the activation barrier. This result is different to similar studies of an earlier generation of molecular motors, which suggests different microscopic mechanisms are in operation in the different generations. Finally, the rate of photochemical isomerization was studied for the series of motors, and those with the largest volume substituents showed the highest photochemical cross section.

Ultrafast Excited State Dynamics in Molecular Motors: Coupling of Motor Length to Medium Viscosity.

Synthetic molecular motors continue to attract great interest due to their ability to transduce energy into nanomechanical motion, the potential to do work and drive systems out-of-equilibrium. Of particular interest are unidirectional rotary molecular motors driven by chemical fuel or light. Probing the mechanistic details of their operation at the single-molecule level is hampered by the diffraction limit, which prevents the collection of dynamic positional information by traditional optical methods. Here, we use defocused wide-field imaging to examine the unidirectional rotation of individual molecular rotary motors on a quartz surface in unprecedented detail. The sequential occupation of nanomechanical states during the UV and heat-induced cycle of rotation are directly imaged in real-time. The approach will undoubtedly prove important in elucidating the mechanistic details and assessing the utility of novel synthetic molecular motors in the future.

Defocused Imaging of UV-Driven Surface-Bound Molecular Motors.

A new unidirectional light-driven molecular motor suitable for host-guest surface inclusion complexes with tris(o-phenylene)cyclotriphosphazene (TPP) was synthesized. The motor molecules formed regular two-dimensional trigonal arrays covering the large facets of disc-shaped TPP nanocrystals. Photochemical and thermal isomerization studies demonstrated that the light-driven rotation of the anchored motors is similar to that observed in solution and is not compromised neither by either the surface confinement or the density of surface coverage (50 vs 100%).

Surface Inclusion of Unidirectional Molecular Motors in Hexagonal Tris(o-phenylene)cyclotriphosphazene.

Light-driven molecular motors derived from chiral overcrowded alkenes are an important class of compounds in which sequential photochemical and thermal rearrangements result in unidirectional rotation of one part of the molecule with respect to another. Here, we employ anion photoelectron spectroscopy to probe the electronic structure and dynamics of a unidirectional molecular rotary motor anion in the gas-phase and quantum chemistry calculations to guide the interpretation of our results. We find that following photoexcitation of the first electronically excited state, the molecule rotates around its axle and some population remains on the excited potential energy surface and some population undergoes internal conversion back to the electronic ground state. These observations are similar to those observed in time-resolved measurements of rotary molecular motors in solution. This work demonstrates the potential of anion photoelectron spectroscopy for studying the electronic structure and dynamics of molecular motors in the gas-phase, provides important benchmarks for theory and improves our fundamental understanding of light-activated molecular rotary motors, which can be used to inform the design of new photoactivated nanoscale devices.

Unravelling the electronic structure and dynamics of an isolated molecular rotary motor in the gas-phase.

A striking feature of living systems is their ability to produce motility by amplification of collective molecular motion from the nanoscale up to macroscopic dimensions. Some of nature's protein motors, such as myosin in muscle tissue, consist of a hierarchical supramolecular assembly of very large proteins, in which mechanical stress induces a coordinated movement. However, artificial molecular muscles have often relied on covalent polymer-based actuators. Here, we describe the macroscopic contractile muscle-like motion of a supramolecular system (comprising 95% water) formed by the hierarchical self-assembly of a photoresponsive amphiphilic molecular motor. The molecular motor first assembles into nanofibres, which further assemble into aligned bundles that make up centimetre-long strings. Irradiation induces rotary motion of the molecular motors, and propagation and accumulation of this motion lead to contraction of the fibres towards the light source. This system supports large-amplitude motion, fast response, precise control over shape, as well as weight-lifting experiments in water and air.

Artificial muscle-like function from hierarchical supramolecular assembly of photoresponsive molecular motors.

The design of a multicomponent system that aims at the direct visualization of a synthetic rotary motor at the single molecule level on surfaces is presented. The synthesis of two functional motors enabling photochemical rotation and fluorescent detection is described. The light-driven molecular motor is found to operate in the presence of a fluorescent tag if a rigid long rod (32 Å) is installed between both photoactive moieties. The photochemical isomerization and subsequent thermal helix inversion steps are confirmed by H NMR and UV-vis absorption spectroscopies. In addition, the tetra-acid functioned motor can be successfully grafted onto amine-coated quartz and it is shown that the light responsive rotary motion on surfaces is preserved.

Design, Synthesis, and Isomerization Studies of Light-Driven Molecular Motors for Single Molecular Imaging.

Control over dynamic functions in larger assemblies is key to many molecular systems, ranging from responsive materials to molecular machines. Here we report a molecular motor that forms bowl-shaped particles in water and how confinement of the molecular motor affects rotary motion. Studying the aggregation process in a broader context, we provide evidence that, in the case of bowl-shaped particles, the structures are not the product of self-assembly, but a direct result of the mixing a good solvent and a (partial) non-solvent and highly independent of the molecular design. Under the influence of the non-solvent, droplets are formed, of which the exterior is hardened due to the increase in the glass transition temperature by the external medium, while the interior of the droplets remains plasticized by the solvent, resulting in the formation of stable bowl-shaped particles with a fluid interior, a glass-like exterior, and a very specific shape: dense spheres with a hole in their side. Applying this to a bulky first-generation molecular motor allowed us to change its isomerization behavior. Furthermore, the motor shows in situ photo-switchable aggregation-induced emission. Strong confinement prohibits the thermal helix inversion step while altering the energy barriers that determine the rotary motion, such that it introduces a reverse trans- cis isomerization upon heating. These studies show a remarkable control of forward and backward rotary motion by simply changing solvent ratios and extent of confinement.

Solvent Mixing To Induce Molecular Motor Aggregation into Bowl-Shaped Particles: Underlying Mechanism, Particle Nature, and Application To Control Motor Behavior.

In living tissues, mechanical stiffness and biological function are intrinsically linked. Alterations in the stiffness of tissues can induce pathological interactions that affect cellular activity and tissue function. Underlying connections between tissue stiffness and disease highlights the importance of accurate quantitative characterizations of soft tissue mechanics, which can improve our understanding of disease and inform therapeutic development. In particular, accurate measurement of lung mechanical properties has been especially challenging due to the anatomical and mechanobiological complexities of the lung. Discrepancies between measured mechanical properties of dissected lung tissue samples and intact lung tissues in vivo has limited the ability to accurately characterize integral lung mechanics. Here, we report a non-destructive vacuum-assisted method to evaluate mechanical properties of soft biomaterials, including intact tissues and hydrogels. Using this approach, we measured elastic moduli of rat lung tissue that varied depending on stress-strain distribution throughout the lung. We also observed that the elastic moduli of enzymatically disrupted lung parenchyma increased by at least 64%. The reported methodology enables assessment of the nonlinear viscoelastic characteristics of intact lungs under normal and abnormal (i.e., injured, diseased) conditions and allows measurement of mechanical properties of tissue-mimetic biomaterials for use in therapeutics or in vitro models. STATEMENT OF SIGNIFICANCE: Accurate quantification of tissue stiffness is critical for understanding mechanisms of disease and developing effective therapeutics. Current modalities to measure tissue stiffness are destructive and preclude accurate assessment of lung mechanical properties, as lung mechanics are determined by complex features of the intact lung. To address the need for alternative methods to assess lung mechanics, we report a non-destructive vacuum-based approach to quantify tissue stiffness. We applied this method to correlate lung tissue mechanics with tissue disruption, and to assess the stiffness of biomaterials. This method can be used to inform the development of tissue-mimetic materials for use in therapeutics and disease models, and could potentially be applied for in-situ evaluation of tissue stiffness as a diagnostic or prognostic tool.

Non-destructive vacuum-assisted measurement of lung elastic modulus.

Injured or diseased airway epithelium due to repeated environmental insults or genetic mutations can lead to a functional decline of the lung and incurable lung diseases. Bioengineered airway tissue constructs can facilitate  investigation of human lung diseases and accelerate the development of effective therapeutics. Here, we report robust tissue manipulation modalities that allow: (i) selective removal of the endogenous epithelium of  cultured airway tissues and (ii) spatially uniform distribution and prolonged cultivation of exogenous cells that are implanted topically onto the denuded airway lumen. Results obtained highlight that our approach to airway tissue manipulation can facilitate controlled removal of the airway epithelium and subsequent homogeneous distribution of newly implanted cells. This study can contribute to the creation of innovative tissue engineering methodologies that can facilitate the treatment of lung diseases, such as cystic fibrosis, primary ciliary dyskinesia, and chronic obstructive pulmonary disease.

Homogeneous Distribution of Exogenous Cells onto De-epithelialized Rat Trachea via Instillation of Cell-Loaded Hydrogel.

Recent synergistic advances in organ-on-chip and tissue engineering technologies offer opportunities to create -grown tissue or organ constructs that can faithfully recapitulate their  counterparts. Such  tissue or organ constructs can be utilized in multiple applications, including rapid drug screening, high-fidelity disease modeling, and precision medicine. Here, we report an imaging-guided bioreactor that allows  monitoring of the lumen of  airway tissues during controlled  tissue manipulation and cultivation of isolated rat trachea. Using this platform, we demonstrated partial removal of the rat tracheal epithelium (, de-epithelialization) without disrupting the underlying subepithelial cells and extracellular matrix. Through different tissue evaluation assays, such as immunofluorescent staining, DNA/protein quantification, and electron beam microscopy, we showed that the epithelium of the tracheal lumen can be effectively removed with negligible disruption in the underlying tissue layers, such as cartilage and blood vessel. Notably, using a custom-built micro-optical imaging device integrated with the bioreactor, the trachea lumen was visualized at the cellular level, and removal of the endogenous epithelium and distribution of locally delivered exogenous cells were demonstrated . Moreover, the de-epithelialized trachea supported on the bioreactor allowed attachment and growth of exogenous cells seeded topically on its denuded tissue surface. Collectively, the results suggest that our imaging-enabled rat trachea bioreactor and localized cell replacement method can facilitate creation of bioengineered  airway tissue that can be used in different biomedical applications.

ArticleTotal : 2	Year
Publication title Cited by 1	2019
Publication title Cited by 2	2022

Article	Year
Molecular stirrers in action. Journal of the American Chemical Society\| PubMed ID: 25254645	2014
Loading of Vesicles into Soft Amphiphilic Nanotubes using Osmosis. Angewandte Chemie (International ed. in English)\| PubMed ID: 26503858	2015
Amphiphilic Molecular Motors for Responsive Aggregation in Water. Journal of the American Chemical Society\| PubMed ID: 26700073	2016
Intramolecular transport of small-molecule cargo in a nanoscale device operated by light. Chemical communications (Cambridge, England)\| PubMed ID: 27068214	2016
End-capping of amphiphilic nanotubes with phospholipid vesicles: impact of the phospholipid on the cap formation and vesicle loading under osmotic conditions. Chemical communications (Cambridge, England)\| PubMed ID: 27709184	2016
Ultrafast Excited State Dynamics in Molecular Motors: Coupling of Motor Length to Medium Viscosity. The journal of physical chemistry. A\| PubMed ID: 28218530	2017
Defocused Imaging of UV-Driven Surface-Bound Molecular Motors. Journal of the American Chemical Society\| PubMed ID: 28497688	2017
Surface Inclusion of Unidirectional Molecular Motors in Hexagonal Tris(o-phenylene)cyclotriphosphazene. Journal of the American Chemical Society\| PubMed ID: 28654753	2017
Unravelling the electronic structure and dynamics of an isolated molecular rotary motor in the gas-phase. Chemical science\| PubMed ID: 28989644	2017
Artificial muscle-like function from hierarchical supramolecular assembly of photoresponsive molecular motors. Nature chemistry\| PubMed ID: 29359757	2018
Design, Synthesis, and Isomerization Studies of Light-Driven Molecular Motors for Single Molecular Imaging. The Journal of organic chemistry\| PubMed ID: 29741383	2018
Solvent Mixing To Induce Molecular Motor Aggregation into Bowl-Shaped Particles: Underlying Mechanism, Particle Nature, and Application To Control Motor Behavior. Journal of the American Chemical Society\| PubMed ID: 29879351	2018
Non-destructive vacuum-assisted measurement of lung elastic modulus. Acta biomaterialia\| PubMed ID: 34192570	2021
Homogeneous Distribution of Exogenous Cells onto De-epithelialized Rat Trachea via Instillation of Cell-Loaded Hydrogel. ACS biomaterials science & engineering\| PubMed ID: 34874712	2022
Imaging-guided bioreactor for de-epithelialization and long-term cultivation of rat trachea. Lab on a chip\| PubMed ID: 35166739	2022

Jiawen Chen

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