Gas-Induced Phase Transition of Dipeptide Supramolecular Assembly

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Metadata	Details
Publication Date	2020-12-08
Journal	CCS Chemistry
Authors	Huimin Xue, Jinbo Fei, Aoli Wu, Xia Xu, Junbai Li
Institutions	University of Chinese Academy of Sciences, Beijing National Laboratory for Molecular Sciences
Citations	1

Abstract

Open AccessCCS ChemistryRESEARCH ARTICLE1 Nov 2021Gas-Induced Phase Transition of Dipeptide Supramolecular Assembly Huimin Xue, Jinbo Fei, Aoli Wu, Xia Xu and Junbai Li Huimin Xue Beijing National Laboratory for Molecular Sciences (BNLMS), CAS Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of Chinese Academy of Sciences, Beijing 100049 , Jinbo Fei *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Beijing National Laboratory for Molecular Sciences (BNLMS), CAS Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of Chinese Academy of Sciences, Beijing 100049 , Aoli Wu Beijing National Laboratory for Molecular Sciences (BNLMS), CAS Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 , Xia Xu Beijing National Laboratory for Molecular Sciences (BNLMS), CAS Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of Chinese Academy of Sciences, Beijing 100049 and Junbai Li *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Beijing National Laboratory for Molecular Sciences (BNLMS), CAS Key Lab of Colloid, Interface and Chemical Thermodynamics, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 University of Chinese Academy of Sciences, Beijing 100049 https://doi.org/10.31635/ccschem.021.202000601 SectionsSupplemental MaterialAboutAbstractPDF ToolsAdd to favoritesTrack Citations ShareFacebookTwitterLinked InEmail The manipulation of supramolecular assembly enables single-component architectures to possess diverse structures and functions. Here, we report directed phase transitions of dipeptide supramolecular gel to crystals with excellent selectivity and tunable mechanical properties. To be specific, lamellar-to-orthorhombic rearrangement of dipeptide molecules in the supramolecular assembly was guided by application of ammonia gas, while lamellar-to-hexagonal realignment was generated upon water vapor exposure of the assembly. Importantly, this crystal phase control originated from distinct gas-mediated reconstitution of hydrogen-bonding interactions, which endowed the dipeptide materials with remarkably modulated stiffness. The selective phase transformation offers a simple and effective platform for self-assembling peptide crystals with diverse long-range-ordered structures from a single gel-state aggregation. This work opens up new perspectives on peptide-based biomaterials via gas-directed hydrogen-bonding chemistry. Download figure Download PowerPoint Introduction Controlled supramolecular assembly offers a promising approach to construct well-defined architectures with tunable structures and functions.1-4 By precisely modulating the self-assembly pathways, supramolecular frameworks with distinct-ordered molecule alignment can be obtained.5-8 Owing to the intrinsic dynamic and adaptive features, supramolecular systems are tailored through subtle variations in molecular arrangements, triggered by external stimuli (light, temperature, pressure, solvents, and chemicals).9-12 In particular, controlled assembly of a single-component building block enables the formation of multiple phases with a rich diversity of intriguing physicochemical properties.13-16 Hence, crystal phase control of ordered supramolecular assemblies can generate new structures and unique functions. However, the process remains a vast challenge under ambient conditions.17-20 The central recognition of Alzheimer’s β-amyloid polypeptide diphenylalanine (FF) motif has attracted much attention owing to the outstanding advantages of identifying the early onset of the disease. This includes its unique structural simplicity and functional versatility features.21,22 Various FF-based architectures, such as vesicles, nanotubes, nanorods, and nanofibers, have been assembled in controlled fashions.23-26 Such molecular assemblies play crucial roles in facilitating the corresponding functions.27-29 For instance, FF materials exhibit tough and flexible properties simultaneously, attributable to the co-existence of laminated organization and hexagonal packing.30 In addition, FF microtubes possess active optical waveguiding properties, associated with the long-range hexagonal alignment of molecules.31 Notably, as an excellent low-molecular-weight gelator, FF can instantly self-assemble into metastable supramolecular gels in a variety of organic solvents.32,33 In previous reports, most gel-crystal phase transitions of FF assemblies were mediated by temperature and liquid solvents.34,35 Nonetheless, gas molecules seem not to have been explored as effective external stimuli. Moreover, for existing FF crystals arising from solution or gel states, the long-range-ordered arrangement of FF molecules is limited to hexagonal packing. Here, we developed a gas-activated approach to achieve selective gel-crystal phase transformations of dipeptide supramolecular assemblies under ambient conditions. As shown in Figure 1, FF readily forms a metastable supramolecular gel via hydrogen-bonding and aromatic stacking interactions in aromatic solvents. With the diffusion of ammonia gas and water vapor into the isotropic supramolecular gel, selective phase transformation was activated to form distinct stable crystal phases through reconstitution of hydrogen-bonding interactions. Consequently, the lamellar arrangement of dipeptide molecules was tuned to orthorhombic (crystal I) and hexagonal (crystal II) realignment, respectively. To the best of our knowledge, this is the first report that has demonstrated the fabrication of orthorhombic-phase dipeptide crystals under ambient conditions. The gas-involved hydrogen-bonding rearrangement chemistry enabled directional evolution of single supramolecular gel into diverse crystals with marked improvement in alignment and remarkably enhanced orientation. Furthermore, the above-mentioned crystal phase control allowed the dipeptide assemblies to exhibit significant tunable mechanical properties. Figure 1 | Schematic illustration of gas-activated phase transitions of dipeptide supramolecular systems through hydrogen-bonding-based rearrangement chemistry. As a result, selective phase transformations of single supramolecular gel into diverse crystal phases are achieved. Oxygen and nitrogen atoms are represented in blue and red, respectively. Download figure Download PowerPoint Experimental Methods Materials FF and 1,1,1,3,3,3-hexafluoro-2-isopropanol (HFIP) were purchased from Sigma-Aldrich (Beijing, China). High-purity ammonia gas (99.999%) was from the Beijing TaiLong Electronics Company (Beijing, China). Toluene, benzene, xylene, ortho-xylene, and styrene were obtained from the Beijing Chemical Reagents Company (Beijing, China). Deionized water was prepared in a three-stage Millipore Milli-Q plus 185-purification system and resistivity >18.2 MΩ cm. All chemicals were used as received unless otherwise stated. Preparation of FF supramolecular gels Precisely 5 mg FF powder was placed in a pre-cleaned bottle with a volume of 3 mL and completely dissolved in HFIP to form FF/HFIP solution with a concentration of 0.4 M. After filtering the solution, 1 mL of filtered toluene was added quickly to generate FF/toluene organogel. Then the FF/HFIP stock solution was filtered through 0.45 μm poly(tetrafluoroethylene) (PTFE). Other FF-based organogels were prepared in the same way, except for the change of solvent. Ammonia gas-induced gel-crystal phase transition of FF supramolecular assembly FF/toluene organogel contained in a 3 mL bottle was transferred into a sealed container rapidly. Next, high-purity ammonia gas was delivered gradually into the container at constant atmospheric pressure to generate a base ammonia environment. After 120 min, the white precipitate formed at the bottom was washed three times via centrifugation (8000 rmp for 6 min) and vortexing with filtered toluene. The precipitate recovered was then sealed or dried under vacuum for further characterization. Water vapor-induced gel-crystal phase transition of FF supramolecular assembly The FF/toluene organogel contained in the 3 mL bottle was transferred quickly into a sealed container. Next, water vapor was delivered into the container continuously through a steam engine at room temperature. After 12 h, the gel became opalescent and had little volume change, and dried for further characterizations. As a comparison, FF/toluene organogel was directly transferred into a sealed container and allowed to stand for 120 min at room temperature without employing ammonia gas or water vapor. No noticeable change in the gel system was noted during this time. Scanning electron microscopy A small piece of gel and a drop of freshly prepared crystal were carefully transferred onto a silicon wafer and dried under a vacuum. Then a thin layer of gold nanoparticles of ∼5 nm was sputtered onto the sample surface. Morphology images were taken using an S-4800 scanning electron microscope (SEM; Hitachi, Japan) with an accelerating voltage of 10 kV. Transmission electron microscopy A dispersed solution of freshly prepared crystal was dropped onto a carbon-coated copper grid and dried in a vacuum. Transmission electron microscopy (TEM) images were taken with a JEM-1011 (JEOL, Japan) under 100 kV. Oscillatory rheology The fresh gel was transferred onto a parallel plate with a diameter of 40 mm and then measured using a Discovery DHR-1 rheometer (TA Instruments, Guangzhou, China). The experiment was performed at 293 K with a frequency of 0.1-100 rad s−1 and a strain of 0.1%. Cross-polarized microscopy A small piece of gel and a drop of freshly prepared crystal were transferred on a glass slide. The cross-polarized microscopy images were taken with a thermal platform microscope (THMS600, BX51; Linkam Scientific, Tadworth, UK). Fluorescence Precisely 5 mg xerogel or dried crystal was ground to compress into a quartz clip with a width of 1 mm. Fluorescence (FL) spectra were recorded using HITACHI F-4500 spectrofluorometer (Shanghai, China) at room temperature. The excitation wavelength was set at 250 nm. Fourier transform infrared Xerogel or dried crystal was mixed with KBr, ground, and compressed into a transparent laminate. The spectroscopic data were recorded using a Fourier transform infrared (FTIR) (Bruker EQUINOX 55/S; Beijing, China). Thermogravimetric analysis Precisely 10 mg of dried samples were put in a crucible. Then thermogravimetric analysis (TGA) curves were obtained on a PerkinElmer Pyris 1 (Chengdu, China) with a heating rate of 10 °C min−1 under nitrogen atmosphere. Powder X-ray diffraction Precisely 5 mg xerogel or dried crystal was ground into the fine powders and pressed on a silicon wafer. Spectra were recorded using a Rigaku D/max-2005 instrument (Beijing, China) equipped with a Cu filter under the following conditions: scan speed, 2° min−1; CuKa radiation, λ = 1.5418 Å. Single-crystal X-ray diffraction A suitable crystal I was selected and mounted on MiTeGen loop (MiTeGen LLC, Ithaca, NY) and flash-frozen in liquid nitrogen. Crystal data were collected at 169.99(10) K on an XtaLAB Synergy R, HyPix diffractometer (Rigaku, Beijing, China). The data were processed using Olex2.2. Then the structure was solved with the ShelXT structure solution program using Intrinsic Phasing36 and refined with the ShelXL refinement package using least-squares minimization.37 The crystallographic data are presented in Supporting Information Table S2. A single-crystal X-ray diffraction of crystal II was obtained using a method described previously.38 The crystallographic data are presented in Supporting Information Table S3. Young’s modulus measurement We used a JPK NanoWizard III atomic force microscope (AFM; JPK Instruments, Berlin, Germany) coupled with an optical microscope for the Young’s modulus measurement. Specifically, the crystals were cast on the surface of a silicon wafer, and the cantilever was moved above the sample with the aid of an optical microscope. For the measurement of crystal I, we adopted the quantitative imaging (QI) mode, which allowed the collection of force curves at the same speed and resolution as normal imaging. Silicon cantilever NCH-50 (Bruker AFM probes) with force constant of 42 N m−1 was used during the experiment. For crystal II measurement, contacting mode and cantilever RTESP-525 with force constant of 200 N m−1 were chosen. The Young’s modulus was obtained and analyzed by fitting the force curves to the Hertz model via the commercial JPK software. All experiments were performed at room temperature. Results and Discussion In a typical experiment, FF (NH2-l-Phe-l-Phe-COOH) supramolecular organogel was prepared by diluting a FF/HFIP solution with toluene at room temperature (25 °C), according to a previous report.34 The inset in Figure 2a shows that the gel is transparent. SEM images (Figures 2a and 2b) revealed that the resulting gel consisted of ultrafine nanofibers. The rheology measurements of the gel showed that the storage modulus (G′) was always greater than the loss modulus (G″), and both of them were very weakly dependent on the oscillatory frequency ( Supporting Information Figure S1), demonstrating the dominant elastic behavior of the gel. After exposure to ammonia gas, the gel transformed into a white precipitate, which accumulated at the bottom of the bottle (Figure 2c, inset), indicating an obvious solid-liquid phase separation. After purification, the precipitate exhibited a transparent shiny surface, as shown in Supporting Information Figure S2a, implying the possibility of crystal formation. Furthermore, the SEM images of Figures 2c and 2d represent crystal I, possessing a regular rectangular platelet-like structure with dimensions of >100 μm in length, ∼20 μm in width, and 2 μm in height. The direction ratio of this highly aligned architecture was >50, which suggested a likely anisotropy along the longitudinal direction. Meanwhile, the TEM image displayed in Supporting Information Figure S3a reveals a rectangular cross-section of a single crystal with a smooth surface. As a comparison, when the gel was exposed to water vapor, there was no apparent solid-liquid phase separation (Figure 2e, inset). Meanwhile, the purified product (crystal II) showed no evident shiny surface ( Supporting Information Figure S2b). In this case, fiber-like crystals with a length >200 μm and a diameter of 150 nm were obtained (Figures 2e and 2f and Supporting Information Figure S3b). These results are in good agreement with a previous study that utilized small liquid water drops.35 Figure 2 | FF organogel and crystal characterization. SEM images of (a and b) gel, (c and d) crystal I mediated by ammonia gas, and (e and f) crystal II induced by water vapor. The insets show the corresponding photographs before and after gel-crystal phase transitions. Download figure Download PowerPoint Additionally, the refractivity of the dipeptide supramolecular assembly was measured before and after phase transition using a cross-polarized microscope. When the relative orientation between the gel and polarizer was rotated, there was no prominent polarization property ( Supporting Information Figure S4a), indicating the isotropic nature of the supramolecular gel. In contrast, as shown in Supporting Information Figures S4b and S4c, crystals I and II exhibited periodical brightness and darkness because light extinction occurred when optical axes of the crystals were parallel to the direction of the objective polarizer.39 These optical changes demonstrated that the FF molecules in the crystals were uniaxially oriented, leading to a strong anisotropy of the refractive index. To obtain in-depth insight into this gel-crystal phase transitions, time-dependent evolution of the supramolecular system was monitored. As shown in Supporting Information Figure S5, after exposure to ammonia gas for 25 min, white solids floated on the gel surface, implying that a phase transition had occurred. Then more solids rapidly appeared in 30 min of reaction time, and an obvious white boundary arose. Along with the increasing time, this boundary of phase transition continued to diffuse with the release of toluene from the gel, resulting in a semi-opaque liquid zone in the middle, which reached the gel bottom by ∼120 min. This might be due to the known gravity effect that makes crystals sink to the bottom successively. These findings suggested three-phase, dynamic sol-gel-crystal boundaries. Especially, Supporting Information Figure S6 reveals the morphologies of the dipeptide assemblies collected from different areas at 120 min. In detail, unformed flakes covered the top, laminated assemblies with short fibers appearing in the middle, with platelet-like crystals precipitate at the bottom. Based on these results, a possible phase transition process could be speculated as follows. First, ammonia gas gradually disassembled the gel network through proton shift, accompanied by the release of the solvent molecule toluene. Next, via rearrangement with the generated ammonia ions, the dissociated FF molecules achieved energetically unfavorable nucleation. Finally, the crystal elongated from the nuclei by following Ostwald’s step rule. For water vapor, it took ∼40 min before the surface of the gel started changing, implying a phase transition initiation ( Supporting Information Figure S7). After 60 min, the top of the gel became opalescent and formed an apparent boundary. Then the boundary continued to move down to the lower region over time, extending with anisotropic speed and became increasingly blurry in the process, which might be attributable to the long fiber structure of the hexagonal crystal. The whole transition process took 12 h, much longer than the case in which the ammonia gas was used. A possible reason might be hypothesized, as follows: When the ammonia gas diffuses into a gel network, proton shift and solvent release co-occur, leading to a faster phase separation and crystal growth. Compared with the case of water vapor, there is no obvious phase separation, and phase transition appears in situ. Moreover, the crystal nanofibers hinder further diffusion of water vapor. Further, after this same gel was allowed to stand for the same period without applying ammonia gas or water vapor, no similar precipitates were formed in the metastable supramolecular system ( Supporting Information Figure S8). We confirmed the versatility of the above approach by choosing FF/benzene, FF/xylene, FF/ortho-xylene, and FF/styrene gels as models for the phase transition studies. The Supporting Information Figure S9 shows that a similar phase transition from a gel to crystal state occurred after ammonia gas stimulation. Similarly, water vapor induced the gel-crystal phase transitions of various FF-based gels ( Supporting Information Figure S10). Moreover, these purified crystals could be preserved at room temperature for longer than 6 months. We sought to unravel the driving forces of gel-crystal phase transitions by investigating the molecular interactions before and after introducing gas molecules into the supramolecular gel. The FL spectra of the gel crystals I and II were recorded, as shown in Figure 3a. After UV excitation at 250 nm, the gel revealed a broad-spectrum emission in the UV region, which peaked at 298 nm. In contrast, blueshift was noted at 285 and 288 nm in crystals I and II, respectively. Meanwhile, each full crystal width at half maximum (FWHM) became narrower. These differences implied stronger π-π attacking interactions between the aromatic residues of FF molecules in the gel, compared with those in the two crystals.40 Figure 3 | (a) Photoluminescence emission spectra (excited at 250 nm), (b) FTIR spectra, (c) TGA curves, and (d) PXRD patterns of the gel, crystals I and II. Download figure Download PowerPoint Moreover, the FTIR spectroscopy displayed in Figure 3b shows the characteristic amide I absorption band (vibration of C=O) at 1670 cm−1 and the amide II absorption band (in-plane vibration of N-H) at 1605 cm−1 in the gel, associated with a hydrogen-bonded antiparallel β-sheet arrangement.41,42 After phase transition, the peak at 1670 cm−1 blueshifted to 1675 and 1685 cm−1 in crystal I and crystal II, while the peak at 1605 cm−1 redshifted to 1578 and 1565 cm−1, respectively. These results indicated the existence of a predominant parallel β-sheet in both which could be to hydrogen-bonding interactions between gas and FF Specifically, a peak appeared at cm−1 in crystal I, which is typical for the vibration of a that of the were not was the of the typical peak from to cm−1 in crystal I, which was and could be to the formation of the associated of in the In contrast, a peak at cm−1 was distinct in the peak of crystal II. The reason might be that from the enhanced hydrogen-bonding interactions between water molecules and the associated interactions water molecules in this crystal. These findings demonstrated the dominant of of hydrogen-bonding interactions to form the two distinct We TGA to the of these dipeptide As revealed in Figure the gel displayed a loss of between and while the two crystals exhibited These results indicated thermal of the dipeptide supramolecular assembly after phase transitions that might be attributable to the loss of water generated by the of the two FF In detail, the rate of crystal I was the compared with that of the gel and crystal II. The possible reason might be the hydrogen-bonding interactions between ammonia and FF the when the temperature was up to the of three assemblies showed a a similar between and as a of the of We measured powder X-ray diffraction patterns to the changes in the molecular arrangement in the dipeptide supramolecular The of the gel (Figure shows two typical at = and corresponding to a of and respectively. This ratio of suggested the lamellar organization of dipeptide molecules in the supramolecular Additionally, as revealed in Supporting Information Figure the of = and peak of indicated a and of the gel. As a comparison, it exhibited orthorhombic rearrangement of the FF molecules in crystal this crystal phase was under ( Supporting Information Table Specifically, the and peak at = showed a of implying a molecular of crystal Supporting Information Figure exhibited a peak at = with an of and of >200 200 compared with that of the gel-state This further the phase and crystal Moreover, compared with the the peak at = remarkably distinct indicating that crystal I had a and anisotropy in the In contrast, the of crystal II induced by water vapor demonstrated a hexagonal with a previous Especially, a of appeared at the peak of = in crystal II. these findings confirmed that ammonia gas and water vapor crucial roles in distinct-ordered structures arising from single gel-state aggregation. To atomic resolution into the rearrangement a single-crystal analysis of crystal I was We that the crystal structure exhibited an orthorhombic with ammonia in the molecular which indicated that ammonia was in the The crystallographic are presented in Supporting Information Table S2. exposure to ammonia gas, proton from FF to resulting in a formation with a ratio of this structure was by a network of aromatic stacking and Specifically, FF molecules into of through between ammonia and FF molecules in an antiparallel hydrogen-bonding network (Figure and three originated from the and from an amide which a of (Figure the hydrogen-bonding interactions for the formation of the elongated along the direction in crystal The aromatic a π-π that as a between hydrogen-bonded (Figure the of crystal I in the direction. The single-crystal of crystal II ( Supporting Information Table was obtained from a previous in this case, there was no molecular change in the dipeptide crystal I we (Figure with diameter of 10 in crystal II. water molecules with structures in the and formed with FF which in the formation of a (Figure FF molecules were further via with a of Å. the aromatic a π-π arrangement was which the of a between the (Figure Figure | Single-crystal structural (a) of FF molecules and in the of crystal I, (b) the corresponding in crystal I, (c) stacking of FF and water molecules in an of crystal II, (d) corresponding in crystal II. The of Young’s modulus of crystal I and crystal II. The a single The show the hydrogen-bonded interactions. The show aromatic interactions. The and nitrogen atoms are represented in and red, respectively. Download figure Download PowerPoint Furthermore, the mechanical of the crystals was measured using AFM and a set of Young’s modulus was analyzed using As shown in Figure Young’s modulus of crystal I was to be This was much lower than and with was as as in As a comparison, crystal II had an Young’s modulus of (Figure with the of FF in previous indicating a of of in the

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DOI: https://doi.org/10.31635/ccschem.020.202000601