多肽分子自组装

源于自然界中广泛存在的蛋白质自组装现象,近年来多肽的自组装逐渐成为材料学和生物医学等领域的研究热点.通过合理调控多肽的分子结构以及改变外界的环境,多肽分子可以利用氢键、疏水性作用、π-π堆积作用等非共价键力自发或触发地自组装形成形态与结构特异的组装体.由于多肽自身具有良好的生物相容性和可控的降解性能,利用多肽自组装技术构建的各种功能性材料在药物控制释放、组织工程支架材料以及生物矿化等领域内有着巨大的应用前景.本文总结了近年来多肽自组装研究的进展,介绍了多肽自组装技术常见的几种结构模型,概括了多肽自组装的机理,并进一步阐述多肽自组装形成的组装体形态及其在材料学和生物医学等领域里的应用.     

The Synthesis of Cystine Peptides by Iodine Oxidation of S-Trityl-cysteine and S-Acetamidomethyl-cysteine Peptides

Previously reported studies of the iodine oxidation of S-trityl-cysteine peptides and S-acetamidomethyl-cysteine peptides, leading directly to cystine peptides, have been extended. Detailed investigations have been made of the reactivities of the S-trityl and the S-acetamidomethyl group towards iodine in various solvents. In chloroform, methylene chloride, trifluoroethanol, and hexafluoroisopropyl alcohol the differences in the reaction rates of the two groups have been found to be extremely large, allowing the selective conversion of the tritylthio groups to disulfides in the presence of the S-acetamidomethyl derivatives. In a second group of solvents, consisting of methanol, acetic acid, dioxane, and mixtures of these solvents with water, simultaneous iodine oxidation of S-trityl- and S-acetamidomethyl-cysteine peptides leads to a preferential combination of these two residues, resulting in predominantly asymmetrical cystine derivatives. - The suitability of the two sulfur-protecting groups in the synthesis of cyclic cystine peptides has been assessed. - Possible reaction mechanisms are discussed. - The scope and limitations of iodine oxidation in peptide synthesis have been studied. The applicability of the method has been demonstrated in the preparation of the open-chain asymmetrical cystine peptide 5 , the protected somatostatin derivative 17 , and the A(1–13) segment 19 of human insulin, previously employed in the total synthesis of this hormone.     

Photoresponsive iodine-bonded liquid crystals based on azopyridine derivatives with a low phase-transition temperature

Halogen bonding interactions in the formation of liquid crystalline phases have been recognised in recent years. Here, we report a novel series of iodine-bonded liquid crystals using 1,2-diiodotetrafluorobenzene (1,2-DITFB) and azopyridine derivatives (AnAzPy), showing a smectic A phase and concurrent photoresponsive behaviour. These were characterised by using a polarising optical microscope, differential scanning calorimetry and UV-vis absorption spectroscopy. The formation of iodine bonding in the complexes was confirmed by X-ray photoelectron spectroscopy and Raman spectroscopy. Importantly, these iodine-bonded complexes demonstrated a low liquid crystal temperature range (30–50°C) among those reported for photoresponsive halogen-bonded liquid crystals. The molar ratio of the iodine-bonded donor and acceptor was 1:1 upon the self-assembly of the supramolecular complex molecule, as indicated by 1D-WAXD experiments of mixed samples of 1,2-DITFB and AnAzPy with different molar ratios. This study offers a new family of photoresponsive halogen-bonded liquid crystals and broadens the potential applications in their associated systems.     

Electrostatic effects on nanofiber formation of self-assembling peptide amphiphiles

Self-assembling peptide amphiphile molecules have been of interest to various tissue engineering studies. These molecules self-assemble into nanofibers which organize into three-dimensional networks to form hydrocolloid systems mimicking the extracellular matrix. The formation of nanofibers is affected by the electrostatic interactions among the peptides. In this work, we studied the effect of charged groups on the peptides on nanofiber formation. The self-assembly process was studied by pH and zeta potential measurements, FT-IR, circular dichroism, rheology, atomic force microscopy, scanning electron microscopy and transmission electron microscopy. The aggregation of the peptides was triggered upon neutralization of the charged residues by pH change or addition of electrolyte or biomacromolecules. Understanding the controlled formation of the hydrocolloid gels composed of peptide amphiphile nanofibers can lead us to develop in situ gel forming bioactive collagen mimetic nanofibers for various tissue engineering studies including bioactive surface coatings.     

Enhanced self-assembly of the 7–12 sequence of amyloid-β peptide by tyrosine bromination

ABSTRACT

Alzheimer’s disease (AD) is a serious neuropathology related to the misfolded assembly state of amyloid-beta (Aβ40 and Aβ42) peptides. It has been demonstrated that protein post-translation modifications (PPTMs) of the more hydrophilic N-term moiety of the Aβ peptide affect its aggregation kinetics and interaction with the environment. Considering that chlorination and bromination are non-canonical PPTMs found in various metabolic pathways and often correlated to inflammatory responses, halogenation of the Y10 of the Aβ N-term could be a putative in vivo modification with implications in the Aβ peptide aggregation propensity. In this framework, we chose as a model system, a short peptide sequence, DSGYEV (i.e. residues 7–12 of the Aβ N-term) and studied its self-assembly behaviour in comparison to its chlorinated and brominated derivatives. Our results show that Y10 halogenation works as a molecular trigger of the peptide self-assembly in solution, promoting the formation of more structured aggregates.     

Supramolecular non-symmetric dimers derived from cholesterol: synthesis and phase transitional properties

Three series of novel dimeric supramolecules, possessing both inter- as well as intramolecular H-bonding, have been prepared and investigated for their thermal properties. They were obtained in excellent yields by condensing cholesteryl ω-(3-hydroxy-4-formylphen-oxy)alkanoates with 4-(n-alkoxy)benzohydrazides, 3,4-bis(n-decyl-oxy)benzohydrazide and 3,4,5-tris(n-decyloxy)benzohydrazide. The influence of increase in the number of terminal n-alkoxy tails from one to three and the length and parity of the flexible spacer on phase transitional behaviour have been thoroughly investigated with the aid of microscopic, calorimetric and X-ray diffraction (XRD) techniques. The results of these complementary studies clearly illustrate the dependence of thermal behaviour of the dimers on these structural factors. The rigid intermolecular association via H-bonds through complementary benzohydrazide component enforce their self-assembly into frustrated and polar smectic phases; H-bond force coupled with the bulkiness of steroid moiety affects the electrical switching property of this fluid polar structure.     

Gel- and Solid-State-Structure of Dialanine and Diphenylalanine Amphiphiles: Importance of C⋅⋅⋅H Interactions in Gelation

To investigate the role of the capping group in the solution and solid-state self-assembly of short peptide amphiphiles, dialanine and diphenylalanine have been linked via the N-terminus to a benzene (phenyl) and 3-naphthyl capping groups using three different methylene linkers; (CH₂)_n, n=0–4 for the benezene and 0, 1 and 2 for the naphthalene capping group. Atomic force microscopy (AFM), oscillatory rheology, circular dichroism (CD), and IR analysis have been employed to understand the properties of these peptide-based hydrogels. Several X-ray structures of these short peptide gelators give useful conformational information regarding packing. A comparison of these solid state structures with their gel state properties yielded greater insights into the process of self-assembly in short peptide gelators, particularly in terms of the important role of C⋅⋅⋅H interactions appear to play in determining if a short aromatic peptide does form a gel or not.     

Bulk and interfacial behaviour of caseinoglycomacropeptide (GMP)

Caseinoglycomacropeptide (GMP) is a hydrophilic glycopeptide released from milk κ-casein by chymosin hydrolysis during cheese making. GMP is thought to be a potential ingredient for specific dietary applications with several health benefits. In this study GMP was characterized at the air–water interface and its behaviour was related with the self-assembly of GMP in solution as affected by pH. This GMP self-assembly was investigated by dynamic light scattering and the interfacial properties were determined by tensiometry and surface dilatational measurements at pH 4, 5 and 7. At pH 5 GMP exhibited higher surface pressure at equilibrium than at pH 7. At pH 4 the behaviour was more complex due to self-assembly close to GMP pI. Dynamic measurement showed that the adsorption/penetration rate constant (K_ads) is facilitated at higher GMP bulk concentrations, while the rate constant of rearrangement (K_r) decreased at higher GMP concentrations which could be attributed to the existence of a steric restriction due to the higher GMP load at the interface. K_r was higher at pH 5 because of lower electrostatic interactions close to the pI. The viscoelastic properties showed a complex behaviour due to the existence of protein–protein interactions depending on the GMP concentration, on the pH of the bulk and on the rates of diffusion, adsorption and rearrangement of GMP at the air–water interface.     

Designer self-assembling peptide materials

Understanding of macromolecular materials at the molecular level is becoming increasingly important for a new generation of nanomaterials for nanobiotechnology and other disciplines, namely, the design, synthesis, and fabrication of nanodevices at the molecular scale from bottom up. Basic engineering principles for microfabrication can be learned through fully grasping the molecular self-assembly and programmed assembly phenomena. Self- and programmed-assembly phenomena are ubiquitous in nature. Two key elements in molecular macrobiological material productions are chemical complementarity and structural compatibility, both of which require weak and non-covalent interactions that bring building blocks together during self-assembly. Significant advances have been made during the 1990s at the interface of materials chemistry and biology. They include the design of helical ribbons, peptide nanofiber scaffolds for three-dimensional cell cultures and tissue engineering, peptide surfactants for solubilizing and stabilizing diverse types of membrane proteins and their complexes, and molecular ink peptides for arbitrary printing and coating surfaces as well as coiled-coil helical peptides for multi-length scale fractal structures. These designer self-assembling peptides have far reaching implications in a broad spectrum of applications in biology, medicine, nanobiotechnology, and nanobiomedical technology, some of which are beyond our current imaginations. [image: see text]     

Thiazole Boron Difluoride Dyes with Large Stokes Shift,Solid State Emission and Room-Temperature Phosphorescence

The small Stokes shift and weak emission in the solid state are two main shortcomings associated with the boron-dipyrromethene (BODIPY) family of dyes. This study presents the design, synthesis and luminescent properties of boron difluoro complexes of 2-aryl-5-alkylamino-4-alkylaminocarbonylthiazoles. These dyes display Stokes shifts (Δλ, 77–101 nm) with quantum yields (ϕ_FL) up to 64.9 and 34.7 % in toluene solution and in solid state, respectively. Some of these compounds exhibit dual fluorescence and room-temperature phosphorescence (RTP) emission properties with modulable phosphorescence quantum yields (ϕ_PL) and lifetime (τ_p up to 251 μs). The presence of intramolecular H-bonds and negligible π-π stacking revealed by X-ray crystal structure might account for the observed large Stokes shift and significant solid-state emission of these fluorophores, while the enhanced spin-orbit coupling (SOC) of iodine and the self-assembly driven by halogen bonding, π-π and C−H^…π interactions could be responsible for the observed RTP of iodine containing phosphors.     

Programming supramolecular peptide materials by modulating the intermediate steps in the complex assembly pathway: Implications for biomedical applications

Self-assembling peptides form a prominent class of supramolecular materials with in general good biocompatibility. To afford better control over the material properties, tremendous progress has been made in studying the supramolecular organization of the peptide assemblies. This knowledge has helped us to understand the correlation between the molecular structure of the peptide building blocks and the properties of the supramolecular products. However, peptide self-assembly consists of a complex pathway rather than a spontaneous thermodynamic process. This implies that the outcome of the self-assembly is critically governed by the assembly pathway. Here, we are going to discuss how peptide self-assembly can be modulated at the intermediate steps in the self-assembly pathway. The focus will be to demonstrate this engineering approach on the example of zero-dimensional/one-dimensional nanostructure selectivity over the β-sheet assembly pathway. In addition, we provide examples of biomedical applications of such steered peptide assemblies in the field of drug delivery and tissue engineering.     

Perfluoroalkyl chains direct novel self-assembly of insulin

The self-assembly of biopharmaceutical peptides into multimeric, nanoscale objects, as well as their disassembly to monomers, is central for their mode of action. Here, we describe a bioorthogonal strategy, using a non-native recognition principle, for control of protein self-assembly based on intermolecular fluorous interactions and demonstrate it for the small protein insulin. Perfluorinated alkyl chains of varying length were attached to desB30 human insulin by acylation of the ε-amine of the side-chain of LysB29. The insulin analogues were formulated with Zn(II) and phenol to form hexamers. The self-segregation of fluorous groups directed the insulin hexamers to self-assemble. The structures of the systems were investigated by circular dichroism (CD) spectroscopy and synchrotron small-angle X-ray scattering. Also, the binding affinity to the insulin receptor was measured. Interestingly, varying the length of the perfluoroalkyl chain provided three different scenarios for self-assembly; the short chains hardly affected the native hexameric structure, the medium-length chains induced fractal-like structures with the insulin hexamer as the fundamental building block, while the longest chains lead to the formation of structures with local cylindrical geometry. This hierarchical self-assembly system, which combines Zn(II) mediated hexamer formation with fluorous interactions, is a promising tool to control the formation of high molecular weight complexes of insulin and potentially other proteins.     

D-periodic collagen-mimetic microfibers

Self-assembling peptides have been previously designed that assemble into macroscopic membranes, nanotapes, and filaments through electrostatic interactions. However, the formation of highly ordered collagen-like fibrils, which display D-periodic features, has yet to be achieved. In this report, we describe for the first time a synthetic peptide system that self-assembles into a fibrous structure with well-defined periodicity that can be visualized by transmission electron microscopy (TEM). Specifically, we designed and synthesized a peptide that utilizes charged amino acids within the ubiquitous Xaa-Yaa-Gly triad sequence to bias the self-assembly into collagen-like homotrimeric helices that are capable of fibrillogenesis with the production of D-periodic microfibers. Potential molecular mechanisms for peptide assembly into triple-helical protomers and their subsequent organization into structurally defined, linear assemblies were explored through molecular dynamics (MD) simulations. The formation of thermodynamically stable complexes was attributed to the presence of strong electrostatic and hydrogen bond interactions at staggered positions along the linear assembly. This unexpected mimicry of native collagen structure using a relatively simple oligopeptide sequence establishes new opportunities for engineering linear assemblies with highly ordered nano- and microscale periodic features. In turn, the capacity to precisely design periodic elements into an assembly that faithfully reproduces these features over large length scales may facilitate the fabrication of ordered two- and three-dimensional fiber networks containing oriented biologically, chemically, or optically active elements.     

Self-Assembly in Natural and Unnatural Systems

Although there are no fundamental factors hindering the development of nanoscale structures, there is a growing realization that “engineering down” approaches, in other words a reduction in the size of structures generated by lithographic techniques below the present lower limit of roughly 1 μm, may become impractical. It has, therefore, become increasingly clear that only by the development of a fundamental understanding of the self-assembly of large-scale biological structures, which exist and function at and beyond the nanoscale, downwards, and the extension of our knowledge regarding the chemical syntheses of small-scale structures upwards, can the gap between the promise and the reality of nanosystems be closed. This kind of construction of nanoscale structures and nanosystems represents the so-called “bottom up” or “engineering up” approach to device fabrication. Significant progress can be made in the development of nanoscience by transferring concepts found in the biological world into the chemical arena. Central to this mission is the development of simple chemical systems capable of instructing their own organization into large aggregates of molecules through their mutual recognition properties. The precise programming of these recognition events, and hence the correct assembly of the growing superstructure, relies on a fundamental understanding and the practical exploitation of non-covalent bonding interactions between and within molecules. The science of supramolecular chemistry—chemistry beyond the molecule in its very broadest sense—has started to bridge the yawning gap between molecular and macro-molecular structures. By utilizing inter-actions as diverse as aromatic π–π stacking and metal–ligand coordination for the information source for assembly processes, chemists have, in the last decade, begun to use biological concepts such as self-assembly to construct nanoscale structures and superstructures with a variety of forms and functions. Here, we provide a flavor of how self-assembly operates in natural systems and can be harnessed in unnatural ones.     

Effect of ionic strength on the self-assembly, morphology and gelation of pH responsive β-sheet tape-forming peptides

Molecular self-assembly is an intrinsic property of proteins central to their biological functionality. One important industrially interesting property is the ability to control and switch on and off self-assembly using a variety of external chemical and physical triggers. Model peptides have been developed with significantly reduced chemical and structural complexity compared to biological proteins. These are ideal systems for exposing the fundamental principles that drive protein-like self-assembly, as well as for establishing in a quantitative manner their structure-function relationship. We investigate simple, short model peptides that adopt a purely β-strand conformation, align in an antiparallel manner and self-assemble in one dimension in solution into long β-sheet nanotapes and higher order aggregates with no other conformation (i.e., helices, turns or random coils) present in the aggregates. These micrometre-long nanostructures gel in solutions at concentrations as low as 0.2% v/v. Their gel-fluid transition has been previously shown to be controlled by pH, temperature, or by mixing with complementary peptides. Here we show the dramatic effect of another chemical trigger, that of physiological-like salt concentration, on the self-assembly, morphology and gelation of a series of systematically designed charged self-assembling tape-forming peptides, each 11 amino acid residues in length, in the pH range of 2-14. This study provides a detailed understanding of the self-assembly of this class of peptides in aqueous solutions of biologically relevant pH and ionic strength. This insight has led to the development of injectable self-assembling peptide lubricants as potential therapeutics for the treatment of early stage knee joint osteoarthritis.     

Solvent-Dependent Nanostructures Based on Active π-Aggregation Induced Emission Enhancement of New Carbazole Derivatives of Triphenylacrylonitrile

In the present study, the carbazole and 2,3,3-triphenylacrylonitrile (TPAN) nanostructures (2-CTPAN and 2,2′-CTPAN) have been designed and synthesized by Pd-catalyzed Sonogashira cross-coupling reaction. CTPAN exhibit aggregation-induced emission enhancement (AIEE) behavior in water with high fluorescence quantum yield. Both the compounds show tunable self-assembly in water as well as in N,N-dimethylformamide (DMF) by extended π–π stacking interactions. CTPAN can be self-assembled into spherical particles in water and the structures of these self-assemblies have been investigated using X-ray diffraction. Interestingly, 2-CTPAN and 2,2′-CTPAN form organogels with a critical gelation concentration (CGC) of 11 and 15 mg mL⁻¹, respectively, in DMF and exhibit acicular and rod shaped morphology, respectively. The single-crystal structure of 2-CTPAN shows that the intermolecular C−H⋅⋅⋅π interactions lock the molecular conformation into a staircase-shaped supramolecular assembly. These AIEE active compounds reveal high water dispersibility, strong yellow fluorescence with high quantum yield, promising photostability and excellent biocompatibility, which make them potential bioimaging agents.     

Heterotypic Supramolecular Hydrogels Formed by Noncovalent Interactions in Inflammasomes

The advance of structural biology has revealed numerous noncovalent interactions between peptide sequences in protein structures, but such information is less explored for developing peptide materials. Here we report the formation of heterotypic peptide hydrogels by the two binding motifs revealed by the structures of an inflammasome. Specifically, conjugating a self-assembling motif to the positively or negatively charged peptide sequence from the ASCPYD filaments of inflammasome produces the solutions of the peptides. The addition of the peptides of the oppositely charged and complementary peptides to the corresponding peptide solution produces the heterotypic hydrogels. Rheology measurement shows that ratios of the complementary peptides affect the viscoelasticity of the resulted hydrogel. Circular dichroism indicates that the addition of the complementary peptides results in electrostatic interactions that modulate self-assembly. Transmission electron microscopy reveals that the ratio of the complementary peptides controls the morphology of the heterotypic peptide assemblies. This work illustrates a rational, biomimetic approach that uses the structural information from the protein data base (PDB) for developing heterotypic peptide materials via self-assembly.     

异氰酸苯酯诱导的类胶原多肽自组装

Synthetic matrices provide powerful tools for dissecting molecular interactions involved in the organization of the extracellular matrix (ECM), establishment of cell axis polarity, and suppression of neoplasticity in pre-cancerous endothelial cells. Collagen is the most abundant protein in extracellular matrix. A de novo approach is essential for the synthesis of collagen matrices which can have a broad impact on the understanding of matrix biology and our capacity to construct safe and medically useful biomaterials. Conventionally, the ECM has been studied by an analytical "top-down" approach, where the individual components of the matrix are first isolated and then characterized to explore their biochemical and functional properties. Since native collagen is difficult to modify and can engender pathogenic and immunological side effects, its application on tissue regeneration is limited. Therefore, we attempted to synthesize artificial collagen directly through small organic molecule recognition. The collagen-like peptides possess various benefits such as being clean, programmable, and easy to modify; therefore, in recent years, they have been used as ideal substrates for the synthesis of collagen nanomaterials. The self-assembly of collagen-like peptides is mainly driven by various non-covalent interactions such as electrostatic attraction, π-π stacking, and metal coordination. This renders a difficulty in the rational design of uniform nanostructures from short synthesized peptides and demands a novel strategy. To date, small organic molecules have been rarely used for the self-assembly of collagen-like peptides. In the present study, we attempted to use the small organic molecules for the combined supramolecular self-assembly of collagen-like peptides. Initially, the collagen-like peptides, (POG)₆ and (POG)₈, synthesized by the solid-phase synthesis technique, were both modified chemically using 4, 4'-methylene bis(phenyl isocyanate) to obtain the collagen-like hybrid peptides, AP₆ and AP₈, respectively. Phenyl isocyanate contributes to the formation of potential weak forces, such as hydrogen bonds and π-π stacking at the N-terminal regions of the collagen-like hybrid peptides. The purity and molecular weight of the collagen-like hybrid peptides were analyzed using analytical high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption ionization time of flight (MALDI-TOF), respectively. The stability of AP₆ and AP₈ triple helices was analyzed by circular dichroism (CD) spectroscopy. The small organic molecule 4, 4'-methylene bis(phenyl isocyanate) promoted the unfolding of (POG)₆ and increased the melting temperature (T_m) of (POG)₈ from 37.7 to 58.8 ℃to form a triple helix. The hydrodynamic radii of collagen-like hybrid peptides were measured by dynamic light scattering (DLS). Atomic force microscopy (AFM) and transmission electron microscopy (TEM) were used to analyze the morphology of the aggregation states. AFM results showed that the collagen-like hybrid peptides, AP₆ and AP₈, formed nanofibers spontaneously. Consistent with the AFM results, TEM showed that the AP₆ and AP₈ collagen-like hybrid peptides also formed nanofiber structures. The formation of stable complexes was attributed to the presence of multiple weak interactions such as hydrogen bonding, π-π stacking, and hydrophobic interactions. In the present study, we demonstrated that the chemical modification of collagen-like polypeptides at the N-terminus via the small organic molecule, 4, 4'-methylene bis(phenyl isocyanate), promoted the intramolecular and intermolecular assembly of collagen-like peptides. A simple and effective strategy has been developed in this study to promote the self-assembly of collagen-like peptides.     

Silk-inspired low-molecular-weight organogelator

A minimal sequence from natural silks, the tetrapeptide GAGA, has been used as inspiration for the design of a new organogelator. Gels were obtained in several organic solvents, and their microscopic aspects were studied by transmission and cryo-scanning electron microscopies (TEM and cryo-SEM). FT-IR spectroscopy, circular dichroism, and wide-angle X-ray diffraction were used to study the self-assembly features of this molecule, and evidence of an antiparallel beta-sheet organization was obtained. Remarkably, the precise secondary structure uniqueness found in Nature was successfully transferred into a small synthetic analogue that self-assembles driven only by noncovalent interactions.