Rotor orientation direction controls geometric curvature and chirality for assemblies of motor amphiphiles in water

Control over geometric curvature and chirality of assemblies in pure aqueous media is key to the design of responsive materials and molecular machines. Here we show how aggregate geometric curvature and chirality of motor amphiphiles could be switched from bicontinuous calabashes to nanoribbons or from vesicles to nanoribbons by modulating rotor orientation direction with dual light/heat stimuli to influence spontaneous curvature in assemblies. The photoisomerization and thermal helix inversion processes of molecular motors have been studied at the molecular level, and the transformation of supramolecular assemblies has been investigated at the microscopic level. The morphological evolution of the calabash‐shaped assembly can be kinetically captured, suggesting that the bicontinuous calabash‐shaped structures are different from the bowl‐shaped aggregates based on solvent‐driven assembly upon the addition of non‐solvent or solvent. The investigation of dual optical/thermal control of rotor orientation can provide a new strategy for tuning the geometric curvature and chirality of nanoassemblies at the nanoscale, arriving ultimately the clusteroluminescence through‐space electronic communication at responsive supramolecular nanosystems.


INTRODUCTION
Engineering the geometric curvature and chirality with molecular precision is one challenge of supramolecular chemistry. Molecular self-assembly into dynamic, responsive, and large supramolecular structures in the pure water phase for precise control of geometric curvature and chirality is a prerequisite for the proper functioning of biological systems. [1] Inspired by the fascinating examples of complex self-assembled structures created by Nature, the design and development of functional molecules and specific or welldefined supramolecular assemblies for control of geometric curvature and chirality with molecular precision in water that are dynamic in nature and responsive to external stimuli has undergone rapid advances in recent years. [2] A key challenge in supramolecular chemistry is to gain full control over the assembly behavior in water, not only under thermodynamic equilibrium but also through molecular synthetic design by introducing handles to control the geometric curvature and chirality of the assemblies via external stimuli, which This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2022 The Authors. Aggregate published by SCUT, AIEI, and John Wiley & Sons Australia, Ltd. may lead to a better understanding of complex and dynamic processes found in Nature. [3] Photoresponsive supramolecular assemblies in pure aqueous media [4] have attracted much attention, in part because material properties highly depend on the geometric curvature and chirality of the supramolecular constituents with the comparison to natural systems, as well as possibilities for biologically relevant applications. [5] Among photoresponsive building blocks, light-driven molecular motors based on overcrowded alkenes [6] are especially unique because their unidirectional rotary process involving four distinct isomerization states can be controlled with two types of external stimuli (i.e., light and heat) with high precision, for the development of their dynamically control properties. [7] Large geometrical transformations of motor molecular containing two chains connected to a firstgeneration molecular motor core can be programmed by changing the two side chains from the remotion of E-isomers to close proximity of Z-isomers upon photoisomerization to dynamically control macroscopic foam properties, [8] control S C H E M E 1 Schematic illustration of the photo-isomerization of motor amphiphiles and the change in chirality and curvature by modulating rotor orientation direction in water. (A) Chirality control. Curvature control: (B) Bicontinuous calabashes, (C) nanoribbons, and (D) vesicles over the magnetic interaction, [9] induce specific aggregation phenomenon. [10] Compared to the symmetric first-generation motors, the second generation motors are composed of an asymmetric lower "stator" fluorene ring linked to an upper "rotor" by the ethylenic "axle" (a double bond in the ground state), [11][12][13] and 360 • unidirectional rotation of the molecular motor by continuous stimulation with light and heat, leading to having four distinct isomers. [13b] Rotation of the rotor relative to the stator around the axis is caused by excited-state photoisomerization. During the thermal helix inversion, the configuration of the stereogenic center causes the rotor to prefer one direction with respect to the stator. [13b] Chiral changes in the rotation of motor molecules enable the reorganization of liquid crystalline films and thus the rotation of microscopic objects. [14,7a] In addition, molecular motors have been successfully applied to the chiral control of helical polymers. [13b] Although there are several reports on first-generation [10b,10c] and second-generation [6b,11b] motor amphiphiles, utilizing light-driven rotor orientation direction to control the geometric curvature and chirality of complex dynamic molecular systems in water poses a significant challenge for second generation motor amphiphiles containing two chains connected to stator ring.
We address the challenge of how the rotor orientation direction of second-generation motor amphiphiles will influence the geometric curvature and chirality of dynamic molecular systems in aqueous media. Herein we report a unique system based on second-generation motor amphiphiles containing two chains connected to stator ring was designed to develop the control of geometric curvature and chirality for the responsive self-assembled systems from bicontinuous calabashes to nanoribbons or from vesicles to nanoribbons by rotor orientation direction with high precision by light and heat stimulation in water (Scheme 1). It appears that rotor orientation direction of motor amphiphiles is the controlling factor in calabash-shaped aggregates of the assembly, which is very different from those bowl-shaped morphologies by the most commonly used method of selective (non-)solvent mixing. [15,16] Although this bowl-shaped morphology is found in connection with a very wide range of molecules: amphiphiles, [15a,17] pseudo-amphiphiles, [18] hydrophobic molecules, [10,19] block copolymers, [15a-15c,20] and many others structures, [21] to our knowledge, this is the first report of the utilization of modulating rotor orientation direction of motor amphiphiles by two orthogonal stimuli (light and heat) as structure-directing factors for the devel-opment of the control of geometric curvature and chirality for the responsive self-assembled systems from bicontinuous calabashes to nanoribbons or from vesicles to nanoribbons in water. Beyond nanoscale self-assembly, this approach could provide the platform for using geometric curvature to control space electronic communication of pure n-electron-based molecular environments within the bicontinuous calabash and vesicle assemblies and, ultimately, clusteroluminescence.

RESULTS AND DISCUSSION
The amphiphilic second-generation molecular motor designed in this work consists of a lower "stator" fluorene ring connected to an upper "rotor" by an ethylene "axle", as well as both the hydrophilic PEG chain with a N,N-dimethylamino moiety and the hydrophobic alkyl chain attached to "stator" ring. The synthesis is summarized in Scheme 2 (see Supporting Information, Figure S1). Fluorenone 7 was prepared from the commercially available dihydroxyfluorenone 4 by the Williamson etherification, and subsequently by functionalization with an ethylene glycol hydrophilic chain to form asym-di-substituted fluorenone 6, followed by protection of the primary alcohol moieties with TBDPS-Cl. After preparation of hydrazone 8 by Wolff-Kishner-Huang reduction, compound 8 was then coupled to the freshly prepared thione 13 via the Barton-Kellogg reaction [13b] to form overcrowded alkene 9. The compound 9 was deprotected and then subjected to a methanesulfonylation reaction to obtain methanesulfonyl functionalized motor 11. Subsequent reaction of motor 11 with dimethylamine gave the corresponding asymmetric amphiphilic molecular motor 1. The two enantiomers and two Z-E isomerism obtained via chiral HPLC, the stable E (stable (S)-(M)-(E) and stable (R)-(P)-(E)), as well as the stable Z (stable (S)-(M)-(Z) and stable (R)-(P)-(Z)), were confirmed by 1 H-1 H COSY, HMQC, HMBC, 2D NOESY, CD experiments, and ECD calculations. (see Supporting Information, Figures S2, S3, and S11-S50).
Since the unidirectionality of the motor is controlled by the relative configuration at a stereogenic center, the photochemical and thermal isomerization processes of the motor amphiphiles solution were investigated based on motor amphiphiles with S-configuration as models. Figure 1A shows the 360 • unidirectional rotary cycle of the photochemical and thermal isomerization processes typical for motor amphiphiles. Rotor rotation of the motor amphiphiles  Figure 1B(a)) in dichloromethane-d 2 was irradiated with 365 nm light at −40 • C for 2 h, and the selective rotation from unstable-E to stable-E ( Figure 1B (b)) by left at room temperature for 3 h via the THI step. Compared to the stable-Z before irradiation ( Figure 1B(a)), the result of the stable-Z after irradiation ( Figure 1B During the whole process of self-assembly, the N,Ndimethylamino species could be protonated by the acidic gas medium via CO 2 bubbling, leading to the formation of cations stable (S)-(M)-(Z) and stable (S)-(M)-(E) (protonated monomers) (Scheme 1), and a successive variation of the hydrophilic-hydrophobic balance and a better dissolution in water, which was confirmed by the increase of UV absorption band intensity with prolonged CO 2 aeration time in Supporting Information Figure S4. The self-assembly of cations stable (S)-(M)-(Z) and stable (S)-(M)-(E) in aqueous media were observed by scanning transmission electron microscopy (STEM) and scanning electron microscope (SEM). In water, cation stable (S)-(M)-(E) forms spherical aggregates (Figure 2A,B), while cation stable (S)-(M)-(Z) shows the formation of different aggregates, as shown by STEM ( Figure 2E,F) and bicontinuous calabash-shaped structures are formed. Bicontinuous calabashes consist of three-dimensional networks of interconnected branched calabashes. In addition, no morphological changes were observed after storing the bicontinuous calabashes or vesicles at room temperature for 24 h in the dark, suggesting that the assemblies were stable.
Under STEM and SEM observation (Figure 2A,B), the cation stable (S)-(M)-(E) was assembled into well-dispersed spherical aggregates with an average diameter of about 130 nm (curvature k = 0.015 nm −1 ). Subsequent dye encapsulation and release experiments could provide evidence for vesicle structure, which could accommodate the cargos within their interior. Calcein as a cargo was used to confirm Interestingly, freeze−thaw cycles and upon bubbling CO 2 in water could activate stable (S)-(M)-(Z) to begin an unexpected formation of bicontinuous calabash-shaped structures of cations stable (S)-(M)-(Z) in aqueous solution ( Figure 2E,F). The cation stable (S)-(M)-(Z) was able to assemble into bicontinuous calabash-shaped structures with a very uniform wall thickness (∼33 nm) and the average diam-eter (∼260 nm) and the mean curvature (∼0.0077 nm −1 ). An attractive phenomenon observed was that the cavities of calabash-shaped structures are interconnected with each other. The connection channel among bicontinuous calabashshaped structures was clearly shown by the STEM images ( Figure 2E) and SEM images ( Figure 3C), and the yellow arrow indicated the connection channel of bicontinuous calabashes. Bicontinuous calabash-shaped morphology has drawn our attention, as it is both very specific in its shape and very rare in its occurrence.
In view of the unique photochemical and thermal stability of motor amphiphiles, we investigated curvature changes upon photochemical and thermal isomerization by STEM and SEM (Figure 2C

F I G U R E 3 SEM images of the morphological evolution process transformation for bicontinuous calabashes (cations stable (S)-(M)-(Z))
bicontinuous calabashes with smaller curvature and larger size disappear completely and only nanoribbon-like structures were observed in both systems, suggesting one curvature of aggregates after irradiation are changed to zero nm −1 . In addition, the structure of nanoribbons was further investigated using atomic force microscopy (AFM). The result demonstrated that these nanoribbons are multilayer structures made of stacked bilayers with a thickness of ∼4.3 nm (Supporting Information Figure S6). The 1 H NMR measurements provide evidences for the photochemical and thermal Z−E isomerization and for the coexistence of stable (S)-(M)-(Z) and stable (S)-(M)-(E), leading to the formation of co-assembly and the curvature change of the aggregates.
The capture of intermediate structures during the assembly process can provide useful information on the understanding of the formation mechanism of bicontinuous calabashes for cation stable (S)-(M)-(Z). The SEM images of the morphological evolution process transformation in bicontinuous calabashes were shown in Figure 3A-F. On the initial course of formation of the bicontinuous calabash-like morphology assemblies, the kippah vesicles as seeds for structure development were formed ( Figure 3A). The cause of the kippah F I G U R E 4 CD spectra of assemblies in aqueous solutions at room temperature. (A) Bicontinuous calabash, (B) vesicle, and (C) nanoribbon. The blue and red solid lines correspond to the CD signals of the assemblies from the motor amphiphiles of the S and R conformations, respectively vesicle formation is postulated to be interplay of the relative flexibility of the vesicle wall, interfacial tension, and pressure gradients. [16e,22] The next process involves upward growth of kippah vesicles, followed by the formation of unclosed vesicle structures ( Figure 3B). Figure 3C shows the calabashshaped immediate morphology based on unclosed vesicle assemblies. At this point, a connection channel between the two compartments could still be seen, and the red arrow indicates the cavity on the calabash-shaped immediate assembly, which is immediate evidence of a calabash-shaped connection channel. Figure 3D-F display calabash-shaped structures with different opening directions, which could furthermore result in the formation of bicontinuous calabashes. The bicontinuous calabashes are different from the bowl-shaped aggregates with a fluid interior and a glass-like shell based on solvent-driven assembly, [10a] which can shrink and swell reversibly to break through the surface upon addition of non-solvent or solvent.
Important features of the motor amphiphiles are two helical structures (M-helical structure and P-helical structure), two Z-E isomerism, a single stereogenic center in the rotor section, an asymmetric stator, and a central carbon-carbon double bond as an axis of rotation. The methyl substituent on the rotor of the motor amphiphiles is a key parameter for strong conformational preference. It was established that the stable-(S)-enantiomer showed a preferred (M)-helicity, while the stable-(R)-enantiomer showed a preferred (P)helicity. [13b,23] Since the molecular motors with a methyl substituent at the stereogenic center have a chiral center, we measured the CD of the bicontinuous calabash, vesicle, and nanoribbon assemblies in aqueous solutions for the stable-E (stable (S)-(M)-(E) and stable (R)-(P)-(E)), as well as the stable-Z (stable (S)-(M)-(Z) and stable (R)-(P)-(Z)). As shown in Figure 4, the blue and red solid lines correspond to the mirror imaged CD signals of the assemblies from the motor amphiphiles of the S and R conformations, respectively, indicating that the supramolecular chirality of the assemblies followed the molecular chirality of the motors.
The chiroptical response behavior of assemblies in aqueous phase was studied by CD and UV-vis spectra (Supporting Information Figure S7). The aqueous solution of assemblies of cations stable (S)-(M)-(Z) and stable (S)-(M)-(E) were irradiated using λ = 365 nm light at 10 • C, respectively. From the CD spectra (Supporting Information Figure S7a,c), the decreasing band at 203, 221, 241, and 273 nm with prolonged irradiation time indicates that the stable to unstable isomerization has occurred in both assemblies. After irradiation with λ = 365 nm light for 7 min, the PSS 365 (photostationary state) was reached (red line). The formation of the unstable isomers showed a negative absorption at 203, 223, 249, and 287 nm, which is consistent with helicity inversion. After subsequent THI at room temperature for 30 min, the positive CD signal was again observed in both cases, in accordance with a consecutive helicity inversion (black line). As shown in the corresponding UV−vis spectra (Supporting Information Figure S7b,d), the aqueous solution of assemblies has a characteristic strong absorption band at 330−450 nm in both cases, and irradiation at 365 nm light at 10 • C resulted in a similar characteristic bathochromic shift as in MeOH solution (Supporting Information Figure S8), as well as a clear isosbestic point at 409 nm, indicating that both motors in assemblies undergo isomerization from the stable to the unstable. The UV absorptions returned to their initial states after the samples were subsequently left in the dark at room temperature for 30 min, which is indicative of the selective conversion of the unstable isomer to the stable isomer by a thermally induced helix inversion. The above phenomenons of the rotary cycle can also be observed by CD and UV-vis spectra in MeOH solution at −20 • C (Supporting Information Figure S8). This spectral change, which is attributed to an increased twist angle of the central olefin, is in accordance with Z-E photoisomerization of the stable to the unstable isomer of the previously studied unfunctionalized molecular motor analog. [12a] The above results suggested that the chiroptical response behavior of assemblies of cations stable (S)-(M)-(Z) and stable (S)-(M)-(E) in aqueous phase could be controlled selectively by external stimuli light and heat. By measuring the speed of the THI at various temperatures, an Eyring plot was constructed (see Supporting Information Figure S9). The Gibbs free energy of activation (Δ ‡ G • ) was calculated as 87.84 ± 2.9 kJ/mol corresponding to a half-life (t 1/2 ) of around 4.51 min at 25 • C.
For further insight into the nuances of molecular stacking, the interaction energy of dimers in aqueous solution (solvent model: IEFPCM) was calculated at the M062X/6-31G(d) level (Table S1), and the geometries of dimers with the strongest interaction energy were displayed in Figure 5. A close inspection of these geometries suggests the existence of π-π stacking between the stators in the Z-dimer, the Edimer, and the Z-E-dimer, but the existence of π-π stacking between the rotors only observed in E-dimer. For the Z-Edimer, the rotor distribution on both sides could reduce spatial repulsion to a greater extent. The observed variation of the aggregates' size, curvature, and shape can be explained by considering spontaneous curvature based on rotor orientation direction of the motor amphiphiles. The stable (S)-(M)-(Z) F I G U R E 5 Optimized structures of (A,B) Z-dimer, (C,D) E-dimer, and (E,F) co-assembly in the aqueous phase at M06-2X/6-31G (d) level of theory F I G U R E 6 Schematic illustration for the formation of bicontinuous calabash, vesicle, and nanoribbon assemblies with smaller angle (θ) consequently prefer the interdigitation to smaller curvature and larger size, while the stable (S)-(M)-(E) with larger angle (θ) for larger curvature and smaller size (shown in Figure 6). It is noteworthy that this mechanism of controlling self-assembly through spontaneous curvature also exists in biology, [24] and materials. [2d,24c,25] Compared with a high curvature of assemblies for the stable (S)-(M)-(E) with larger angle (θ), the resulting decrease in spontaneous releases the strain, leading to the formation of the bicontinuous calabash-like morphology assemblies to decrease the total interfacial area as illustrated in Figure 6. Upon the irradiated and subsequently heated sample, the stable (S)-(M)-(E) is co-assembled with the stable (S)-(M)-(Z), the interface between the hydrophilic and hydrophobic parts changes from a curved interface to a flat one. In light of the above, we conclude that rotor orientation direction of motor amphiphiles is key for control over the geometric curvature and chirality of assemblies.
Based on the change of geometric curvature of assemblies by modulating rotor orientation direction of motor amphiphiles, the systems could provide the possibility to control space electronic communication of pure n-electron-based molecular environments in the assemblies with different geometric curvature (bicontinuous calabash, vesicle, and nanoribbon). Certain pure n-electron-based small molecules, such as poly(ethylene glycol) (PEG) have been reported to have intrinsic fluorescence in their aggregates or solidstate forms [26] and may also show clusterization-triggered emission (clusteroluminescence). Tang et al. put forward a comprehensive theory of the through-space conjugation (TSC) for these chromophores. [27] Based on the above motor amphiphiles, the curvature effects on the contribution to the clusteroluminescence through-space electronic communication by O⋯O, N⋯N short contacts were evaluated by the modulation of rotor orientation direction of motor amphiphiles. The fluorescence properties of motor amphiphiles (cations stable (S)-(M)-(Z) and cations stable (S)-(M)-(E)) were investigated in aqueous solution. The λ max-em of ethylene glycol group is 398 and 493 nm under the 240 and 380 nm excitation for cations stable (S)-(M)-(Z), respectively. The results showed that the emission wavelengths of the motor amphiphiles are excitation-dependent, that is, excitation at longer wavelengths leads to a redshifted emission (Supporting Information Figure S10). Most clusteroluminogens, if not all, exhibit excitation-dependent emission properties. [28] As shown in Figure 7, the emission spectra of cations stable (S)-(M)-(Z) and cations stable (S)-(M)-(E) show a single peak at approximately 455 nm at a low concentration when excited at 340 nm. Meanwhile, we observed that their emission intensities are enhanced with increasing concentration for cations stable (S)-(M)-(Z) and cations stable (S)-(M)-(E) in aqueous solution, and that their emission intensities and wavelengths are size-dependent for both systems. The result showed that larger-sized aggregates with smaller curvature could emit stronger intensity emission at longer wavelengths at the same concentration, which was ascribed to the modulation of rotor orientation direction of motor amphiphiles reducing the curvature of aggregates thus increasing the interactions between poly(ethylene glycol) groups. All these results suggest that one of the main causes of the red-shifted clusteroluminescence with stronger intensity is the intra-/intermolecular interactions among neighboring isolated ethylene glycol groups. Therefore, the same molecule but at locations of different underlying curvature can exhibit different clusteroluminescence properties.

CONCLUSION
Here we demonstrate the control of geometric curvature and chirality of assemblies in aqueous media by modulating rotor orientation direction of dual light/heat stimuli-responsive motor amphiphiles. Beyond nanoscale self-assembly, the key conceptual advance of this approach is the use of geometric curvature to control space electronic communication of pure n-electron-based molecular environments within the bicontinuous calabash, vesicle and nanoribbon assemblies and, ultimately, clusteroluminescence. In this way, engineering the geometric curvature and chirality with molecular precision can provide a platform that allows the same molecule but at locations of different underlying curvature to modulate their clusteroluminescence properties through space electronic communication which can be extended to π-π*, n-π*, n-σ*, hydrogen bonding and other weak interactions.

C O N F L I C T O F I N T E R E S T
The authors declare no conflict of interest.

A U T H O R C O N T R I B U T I O N S
Yun-Han Yang contributed to investigation, methodology, synthesis, theoretical calculation, and writing-original draft. Yang Qin and Yang Zhang contributed to investigation. Ling Zhang contributed to conceptualization, supervision, and writing-review and editing. All authors approve the publication of the manuscript in the current version.

D ATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are available from the corresponding author upon reasonable request.