Stable Helical Conformation Research Articles

Thermoresponsive Helical Dendronized Poly(phenylacetylene)s: Remarkable Stabilization of Their Helicity via Photo-Dimerization of the Dendritic Pendants.

Dynamic helical polymers can change their helicity according to external stimuli due to the low helix-inversion barriers, while helicity stabilization for polymers is important for applications in chiral recognition or chiral separations. Here, we present a convenient methodology to stabilize dynamic helical conformations of polymers through intramolecular cross-linking. Thermoresponsive dendronized poly(phenylacetylene)s (PPAs) carrying 3-fold dendritic oligoethylene glycol pendants containing cinnamate moieties were synthesized. These polymers exhibit typical features of dynamic helical structures in different solvents, that is, racemic contracted conformations in less polar organic solvents and predominantly one-handed stretched helical conformations in highly polar solvents. This dynamic helicity can be enhanced through selective solvation by increasing the polarity of the organic solvents or simply via their thermally mediated dehydration in water. However, through photocycloaddition of the cinnamate moieties between the neighboring pendants via UV irradiation, these dendronized PPAs adopt stable helical conformations either below or above their phase transition temperatures in water, and their helical conformations can even be retained in less polar organic solvents. Spectroscopic and atomic force microscopy measurements demonstrate that photocycloaddition between the cinnamate moieties occurs on the individual molecular level, and this is found to be helpful in restraining the photodegradation of the PPA backbones. Molecular dynamics simulations reveal that the spatial orientation of the pendants along the rigid polyene backbone is crucial for the photodimerization of cinnamates within one helix pitch.

Helical polyether-immobilized chiral aza-bis(oxazolines): Synthesis and synergistic effect on the enantioselectivity of Zn-catalyzed Henry reaction

Novel polyether-immobilized aza-bis(oxazolines) ligands derived from optically pure epichlorohydrin and aza(bisoxazolines) were prepared by anionic polymerization. Through chiral optical studies, it is determined that the chiral polyether ligands can maintain stable helical conformation in the solvent, with controllable molecular weights and low polydispersity index of 1.02–1.06. Asymmetric Henry reactions of aldehydes were performed to investigate the relationship between the catalytic activity and the helical conformation of these polyethers, gave the adducts in up to 97 % yield with high enantiomeric excess(98 % ee). In particular, there is a synergistic effect between the helical chirality of the polyether backbone and the point chirality of the pendant aza-bis(oxazolines) in the asymmetric Herny reaction. Helical polyether-immobilized chiral aza-bis(oxazolines) ligands can be easily recycled at least 10 times without losing their enantioselectivity. It was also found by MALDI-TOF-MS that zinc(II) combined with polyether probably result in a double spiral species in the presence of more than 2.5 mol% Zn(OAc)2, which lead to negative effect on catalytic performance of the polyether.

Homochiral versus Heterochiral Dimeric Helical Foldamer Bundles: Chlorinated-Solvent-Dependent Self-Sorting.

Aromatic oligoamide sequences programmed to fold into stable helical conformations were designed to display a linear array of hydrogen-bond donors and acceptors at their surface. Sequences were prepared by solid-phase synthesis. Solution 1 H NMR spectroscopic studies and solid-state crystallographic structures demonstrated the formation of stable hydrogen-bond-mediated dimeric helix bundles that could be either heterochiral (with a P and an M helix) or homochiral (with two P or two M helices). Formation of the hetero- or homochiral dimers could be driven quantitatively using different chlorinated solvents-exemplifying a remarkable case of either social or narcissistic chiral self-sorting or upon imposing absolute handedness to the helices to forbid PM species.

Reversible conformational switching of a photo-responsive ortho-azobenzene/2,6-pyridyldicarboxamide heterofoldamer.

We report on a convenient synthetic route to rapidly access a new photo-responsive ortho-azobenzene/2,6-pyridyldicarboxamide heterofoldamer. The adoption of a stable helical conformation has been established for this scaffold in both the solid state and in solution using single crystal X-ray diffraction and circular dichroism (CD) spectroscopy respectively. Reversible control over the stimuli-driven structural re-ordering of the supramolecular scaffold, from a stable helical conformation under non-irradiative conditions, to a less well-ordered state under irradiative conditions, has been identified. The robust nature of the responsive, conformational, molecular switching behaviour has been determined using UV/Vis, 1H NMR and CD spectroscopy. Minimal loss in the efficiency of the stimuli-driven, structural re-ordering processes of the foldamer scaffold is observed, even upon multiple cyclic treatments with irradiative/non-irradiative conditions.

Single-helical formyl β-glucan effectively deliver CpG DNA with poly(dA) to macrophages for enhanced vaccine effects

Single helical β-glucan is a one-dimensional host that can form a hybrid helix with DNAs/RNAs as delivery systems. However, unmodified β-glucan has a gelling tendency and a single helical conformation is challenging to obtain. Therefore, in this study, we developed a β-glucan formyl derivative with stable single helical conformation and no gelling tendency. Circular dichroism studies found that the formyl-β-glucan could form a hybrid helix with DNA CpG-poly(dA). The hybrid helix delivery system showed improved activation on antigen-presenting cells, thereby upregulating the mRNA and protein levels of inflammatory factors, and had an immune-enhancing effect on ovalbumin (OVA) immunized mice. These results indicate that formyl-β-glucan can be developed as a non-cationic supramolecular DNA delivery platform with low toxicity and high efficiency.

Synthesis of Optically Active Helical Polycarbenes through Helix-Sense-Selective Polymerization Strategy and Their Application in Chiral Separation.

In this work, helical polycarbenes with optical activity were designed and facilely synthesized through the helix-sense-selective polymerization (HSSP) of the diazoacetate monomer with a dimethylbenzyl ester pendant catalyzed by π-allylPdCl with chiral phosphine ligands at room temperature. The polymerization was carried out in a living and controlled style, and a range of helical polycarbenes with the desired number-average molecular weights and narrow molecular weight distributions were obtained. Circular dichroism and UV-vis analyses revealed that these polycarbenes exhibited a stable helical conformation with a preferred handedness, and their helical directions were dependent on the chirality of the chiral phosphine ligands. Further studies showed that the helical conformation of the obtained polycarbenes was from the polymeric backbone rather than the intermolecular aggregation in the solutions. Moreover, the prepared, optically active, helical polycarbenes possessed excellent enantioselective crystallization ability for threonine racemates. The enantiomeric excess (e.e.) of the induced crystals could be up to 83% via utilizing the prepared helical polycarbenes as a chiral separation agent.

Stability Enhancement of a π-Stacked Helical Structure Using Substituents of an Amino Acid Side Chain: Helix Formation via a Nucleation-Elongation Mechanism.

Molecular design involving the incorporation of an α-amino acid residue into the side chain or main chain of a polymer is often used to stabilize artificial molecular architectures through intramolecular hydrogen bonding. However, this molecular design strategy rarely considers the importance of interactions between substituents at the α-position of amino acid moieties, as found in nature. Herein, we report the synthesis of a novel series of π-stacked helical poly(quinolylene-2,3-methylene) with amino acid derivatives bearing different substituents at the α-position. We found that the thermal stability of π-stacked helical poly(quinolylene-2,3-methylene) is significantly improved by packing the substituents in the empty spaces between the side chains. In particular, when a bulky cyclohexyl alanine derivative was used as the side chain, the π-stacked helical structure maintained its stability even in dimethylsulfoxide, a hydrogen bond competitor. The stabilization of the π-stacked structure by the amino acid substituents resulted in a unique polymerization behavior involving nucleation-elongation steps. In the case of derivatives with leucine and cyclohexyl alanine, which form stable π-stacked helical structures, metastable structures with entangled main chains were formed in the initial polymerization stage. These structures subsequently underwent an irreversible structural change to achieve a thermodynamically stable helical π-stacked conformation as a nucleus for subsequent polymerization. Thereafter, the polymerization reaction proceeded with the elongation of the π-stacked helical structure. Differences in the stability of these systems indicated that the amino acid substituents on the side chains determine the most thermodynamically stable π-stacked helical structure.

Structural Dissection of the First Events Following Membrane Binding of the Islet Amyloid Polypeptide

The islet amyloid polypeptide (IAPP) is the main constituent of the amyloid fibrils found in the pancreas of type 2 diabetes patients. The aggregation of IAPP is known to cause cell death, where the cell membrane plays a dual role: being a catalyst of IAPP aggregation and being the target of IAPP toxicity. Using ATR-FTIR spectroscopy, transmission electron microscopy, and molecular dynamics simulations we investigate the very first molecular steps following IAPP binding to a lipid membrane. In particular, we assess the combined effects of the charge state of amino-acid residue 18 and the IAPP-membrane interactions on the structures of monomeric and aggregated IAPP. Distinct IAPP-membrane interaction modes for the various IAPP variants are revealed. Membrane binding causes IAPP to fold into an amphipathic α-helix, which in the case of H18K-, and H18R-IAPP readily moves beyond the headgroup region. For all IAPP variants but H18E-IAPP, the membrane-bound helix is an intermediate on the way to amyloid aggregation, while H18E-IAPP remains in a stable helical conformation. The fibrillar aggregates of wild-type IAPP and H18K-IAPP are dominated by an antiparallel β-sheet conformation, while H18R- and H18A-IAPP exhibit both antiparallel and parallel β-sheets as well as amorphous aggregates. Our results emphasize the decisive role of residue 18 for the structure and membrane interaction of IAPP. This residue is thus a good therapeutic target for destabilizing membrane-bound IAPP fibrils to inhibit their toxic actions.

Extracellular Secretion and Simple Purification of Bacterial Collagen from Escherichia coli

Because of structural similarities with type-I animal collagen, recombinant bacterial collagen-like proteins have been progressively used as a source of collagen for biomaterial applications. However, the intracellular expression combined with current costly and time-consuming chromatography methods for purification makes the large-scale production of recombinant bacterial collagen challenging. Here, we report the use of an adapted secretion pathway, used natively byEscherichia colito secrete curli fibers, for extracellular secretion of the bacterial collagen. We confirmed that a considerable fraction of expressed collagen (∼70%) is being secreted freely into the extracellular medium, with an initial purity of ∼50% in the crude culture supernatant. To simplify the purification of extracellular collagen, we avoided cell lysis and used cross-flow filtration or acid precipitation to concentrate the voluminous supernatant and separate the collagen from impurities. We confirmed that the secreted collagen forms triple helical structures, using Sirius Red staining and circular dichroism. We also detected collagen biomarkers via Raman spectroscopy, further supporting that the recombinant collagen forms a stable triple helical conformation. We further studied the effect of the isolation methods on the morphology and secondary structure, concluding that the final collagen structure is process-dependent. Overall, we show that the curli secretion system can be adapted for extracellular secretion of the bacterial collagen, eliminating the need for cell lysis, which simplifies the collagen isolation process and enables a simple cost-effective method with potential for scale-up.

Confined crystallization and chain conformational change in electrospun poly(ethylene oxide) nanofibers

This study focused on understanding the confined crystallization of poly(ethylene oxide) (PEO) in electrospun nanofibers. The effect of thermal treatment on PEO crystallization was also studied. An electrospinning process with a rotating drum collector enabled stretching and aligning of polymer chains, which resulted in the formation of a planar zigzag conformation. The confined environment in nanofibers facilitated an ordered crystal arrangement, resulting in an increase in the degree of crystallinity with a decrease in the fiber diameter. By contrast, large fibers extended solvent evaporation, which resulted in large crystallite sizes. The confined geometry and mechanical force provided by a rotating collector induced the preferred crystal orientation parallel to the fiber axis. Upon thermal annealing, the stretched PEO chains relaxed, resulting in a change from the metastable zigzag conformation to a stable helical conformation. Thermal treatment monotonically increased the melting temperature, degree of crystallinity, and crystallite size of PEO nanofibers regardless of the fiber diameter but did not influence the orientation of the organized crystals. The confinement effect on the chain conformation and crystallization behavior of poly(ethylene oxide) (PEO) in electrospun nanofibers was investigated. The PEO chains in electrospun nanofibers were tightly packed and exhibited a zigzag conformation. Increasing the fiber diameter reduced the packing of polymer chains, resulting in increases in the melting temperature and crystallite size. Thermal treatment relaxed the stretched chains, which induced a change from the metastable zigzag conformation to a stable helical conformation. Additionally, the melting temperature, degree of crystallinity, and crystallite size increased with increasing annealing temperature.

CryoET structures of immature HIV Gag reveal six-helix bundle

Gag is the HIV structural precursor protein which is cleaved by viral protease to produce mature infectious viruses. Gag is a polyprotein composed of MA (matrix), CA (capsid), SP1, NC (nucleocapsid), SP2 and p6 domains. SP1, together with the last eight residues of CA, have been hypothesized to form a six-helix bundle responsible for the higher-order multimerization of Gag necessary for HIV particle assembly. However, the structure of the complete six-helix bundle has been elusive. Here, we determined the structures of both Gag in vitro assemblies and Gag viral-like particles (VLPs) to 4.2 Å and 4.5 Å resolutions using cryo-electron tomography and subtomogram averaging by emClarity. A single amino acid mutation (T8I) in SP1 stabilizes the six-helix bundle, allowing to discern the entire CA-SP1 helix connecting to the NC domain. These structures provide a blueprint for future development of small molecule inhibitors that can lock SP1 in a stable helical conformation, interfere with virus maturation, and thus block HIV-1 infection.

Native-Based Dissipative Particle Dynamics Approach for α-Helical Folding.

We developed a dissipative particle dynamics (DPD) approach that captures polyalanine folding into a stable helical conformation. Within the proposed native-based approach, the DPD parameters are derived based on the contact map constructed from the molecular dynamics (MD) simulations. We show that the proposed approach reproduces the folding of polypeptides of various lengths, including bundle formation for sufficiently long polypeptides. The proposed approach also allows one to capture the folding of the helical segments of the lysozyme. With further development of computationally efficient native-based DPD approaches for folding, modeling of a range of biomaterials incorporating α-helical segments could be extended to time and length scales far beyond those accessible in molecular dynamics simulations.

Synthesis of modifiable photo-responsive polypeptides bearing allyloxyazobenzene side-chains

A photo-responsive and modifiable polypeptide with stable helical conformation was synthesized. The self-assembly and liquid crystalline phase structure were subsequently studied.

Synthesis and enantioseparation of proline-derived helical polyacetylenes as chiral stationary phases for HPLC

Proline-derived aliphatically substituted polyacetylenes with stable helical conformations exhibit an excellent enantioseparation ability as chiral stationary phases of HPLC.

Synthetic approach for optically active polymers through the combination of asymmetric chirogenic polymerization and postpolymerization modification

A new type of asymmetric chirogenic polymerization by asymmetric allylic substitution catalyzed by planar–chiral ruthenium complexes was designed. The polymerization systems function in a highly stereoselective manner to afford optically active polymers with high selectivity. The asymmetric carbon in the main chain is precisely controlled. Each monomer unit of the polymer has a potentially reactive terminal olefin, which can be used for further transformations. Optically active polymers bearing chiral cyclic architecture were prepared by a combination of asymmetric allylic substitution and ring-closing metathesis reaction employing the terminal olefin of the side chains. Additionally, the efficient introduction of a wide range of substituents into the side chain of the optically active polymer without any racemization has been made possible by using the thiol–ene reaction. Poly-N-alkoxyamides obtained by our asymmetric polymerization can be transformed into nonnatural polypeptides containing an aromatic ring on the peptide backbone, called a poly “arylopeptide”, through reductive cleavage of the N–O bond in N-alkoxyamide. The resulting polymer adopts a one-handed stable helical conformation in solution. A new type of asymmetric chirogenic polymerization by asymmetric allylic substitution catalyzed by planar–chiral ruthenium complexes was designed. The polymerization systems function in a highly stereoselective manner. The asymmetric carbon in the main chain is precisely controlled. Each monomer unit of the polymer has a potentially reactive terminal olefin, which can be used for further transformations, ring-closing metathesis and thiol–ene reaction. Additionally, the resulting polymer can be transformed poly “arylopeptide”, through reductive cleavage of the N–O bond in N-alkoxyamide, which adopts a one-handed stable helical conformation in solution.

Immunomodulatory, insulinotropic, and cytotoxic activities of phylloseptins and plasticin-TR from the Trinidanian leaf frog Phyllomedusa trinitatis.

The aim of the study was to determine the in vitro immunomodulatory, cytotoxic, and insulin-releasing activities of seven phylloseptin-TR peptides and plasticin-TR, first isolated from the frog Phyllomedusa trinitatis. The most cationic peptides, phylloseptin-1.1TR and phylloseptin-3.1TR, showed greatest cytotoxic potency against A549, MDA-MB231, and HT-29 human tumor-derived cells and against mouse erythrocytes. Phylloseptin-4TR was the most hydrophobic and the most effective peptide at inhibiting production of the proinflammatory cytokines TNF-α and IL-1β by mouse peritoneal cells but was without effect on production of the antiinflammatory cytokine IL-10. Phylloseptin-2.1TR and phylloseptin-3.3TR were the most effective at stimulating the production of IL-10. The noncytotoxic peptide, plasticin-TR, inhibited production of TNF-α and IL-1β but was without effect on IL-10 production. The results of CD spectroscopy suggest that the different properties of plasticin-TR compared with the immunostimulatory activities of the previously characterized plasticin-L1 from Leptodactylus laticeps may arise from greater ability of plasticin-TR to oligomerize and adopt a stable helical conformation in a membrane-mimetic environment. All peptides stimulated release of insulin from BRIN-BD11 rat clonal β cells with phylloseptin-3.2TR being the most potent and effective and phylloseptin-2.1TR the least effective suggesting that insulinotropic potency correlates inversely with helicity. The study has provided insight into structure-activity relationships among the phylloseptins. The combination of immunomodulatory and insulinotropic activities together with low cytotoxicity suggests that phylloseptin-3.3TR and plasticin-TR may represent templates for the development of agents for use in antiinflammatory and type 2 diabetes therapies.

A Facile Synthetic Route to Amphiphilic Poly(Meta-Phenylene Ethynylene) and Poly(Meta-Phenylene Ethynylene)-Block-Polyisocyanide Using a Single Catalyst.

An optically active, amphiphilic meta-phenylene ethynylene (m-PE) bearing a chiral amide pendant was designed and synthesized. Living polymerization of m-PE using alkyne-Pd(II) as the initiator afforded well-defined poly(meta-phenylene ethynylene) (m-PPE). These m-PPEs were found to have a stable helical conformation in THF, 1,4-dioxane, and CH3CN and showed split Cotton effects over the range of 245–400 nm. The positive first Cotton effect was observed at a wavelength of approximately 308 nm, and the negative second Cotton effect was observed at a wavelength of approximately 289 nm. The m-PPEs exhibited helical conformational changes in different mixed solvents and showed effective solvent-dependent helix inversion in CHCl3/THF solutions. The sign of the Cotton effect of m-PPE was inverted at 25 °C by varying the mixing ratio of THF and CHCl3. Finally, amphiphilic poly(meta-phenylene ethynylene)-block-polyisocyanide containing hydrophilic PPE and hydrophobic PPI segments were facilely prepared using Pd(II)-terminated m-PPE as the macroinitiator. This block copolymer can self-assemble into well-defined spherical nanostructures in a selective THF/CH3OH solution. This efficient polymerization will open up enormous opportunities for the preparation of functional amphiphilic block copolymers in a wide variety of fields.

Matched and Mismatched Phenomena in the Helix Orientation Bias Induced by Chiral Appendages at Multiple Positions of Indolocarbazole-Pyridine Hybrid Foldamers.

A series of indolocarbazole-pyridine (IP) hybrid foldamers containing chiral residues at multiple different positions were prepared to reveal the matched and mismatched phenomena of local stereocenters on the induction of helical bias. These foldamers adopted stable helical conformations, thus affording well-resolved separate sets of 1H NMR signals for right- ( P) and left-handed ( M) helices in water saturated organic solvents such as toluene and dichloromethane. The ratios of P- and M-helices were determined by integrating the 1H NMR signals, in combination with the molar circular dichroism (Δε) and optical rotation ([α]D) values. The degree of helical bias was larger in the IP foldamer bearing chiral residues at the termini relative to those at the pyridine side chains, but the preferred helix orientation was opposite to each other. Foldamers 5( SS)t( SSS)py and 6( RR)t( SSS)py with chiral residues at five different positions demonstrated the matched and mismatched phenomena of local stereocenters in 6( RR)t( SSS)py and 5( SS)t( SSS)py, respectively.

Novel bioactive peptides from PD-L1/2, a type 1 ribosome inactivating protein from Phytolacca dioica L. Evaluation of their antimicrobial properties and anti-biofilm activities

Antimicrobial peptides, also called Host Defence Peptides (HDPs), are effectors of innate immune response found in all living organisms. In a previous report, we have identified by chemical fragmentation, and characterized the first cryptic antimicrobial peptide in PD-L4, a type 1 ribosome inactivating protein (RIP) from leaves of Phytolacca dioica L. We applied a recently developed bioinformatic approach to a further member of the differently expressed pool of type 1 RIPs from P. dioica (PD-L1/2), and identified two novel putative cryptic HDPs in its N-terminal domain. These two peptides, here named IKY31 and IKY23, exhibit antibacterial activities against planktonic bacterial cells and, interestingly, significant anti-biofilm properties against two Gram-negative strains. Here, we describe that PD-L1/2 derived peptides are able to induce a strong dose-dependent reduction in biofilm biomass, affect biofilm thickness and, in the case of IKY31, interfere with cell-to-cell adhesion, likely by affecting biofilm structural components. In addition to these findings, we found that both PD-L1/2 derived peptides are able to assume stable helical conformations in the presence of membrane mimicking agents (SDS and TFE) and intriguingly beta structures when incubated with extracellular bacterial wall components (LPS and alginate). Overall, the data collected in this work provide further evidence of the importance of cryptic peptides derived from type 1 RIPs in host/pathogen interactions, especially under pathophysiological conditions induced by biofilm forming bacteria. This suggests a new possible role of RIPs as precursors of antimicrobial and anti-biofilm agents, likely released upon defensive proteolytic processes, which may be involved in plant homeostasis.

Binding modes of Bcl-2 homology 3 (BH3) peptides with anti-apoptotic protein A1 and redesign of peptide inhibitors: a computational study

The interaction between protein and peptide ligand is a challenging problem in molecular biology and drug design. The binding of the Bcl-2 homology 3 (BH3) peptide to the anti-apoptotic protein A1 was revealed as a critical step in the regulation of apoptosis. These BH3 peptides hold high structural similarity, but are diverse in their regulation abilities. Based on molecular simulations and MM-P(G)BSA methods, this work presented a detailed analysis on binding mechanism of the BH3 peptides derived from PUMA and BMF. Residue-level energy decomposition showed that the core regions of BH3 peptides maintain in stable helical conformations and the four conserved hydrophobic residues together with an invariant aspartic acid contribute the major driving force for binding, whereas their two terminal segments exhibit obvious flexibility and various binding modes. Such kind of behavior was suggested as the reason for binding diversity and selectivity of BH3 peptides. As a further step, several BH3-mimetic peptides have been redesigned by computational mutation. Those new peptides showed not only stronger affinities when binding to protein A1, as well transferable binding patterns at some specific positions. A long-range coupling effect was disclosed for BH3 peptides, side-chain orientation and binding contribution of terminal residues were even affected by mutations at large sequence interval. Overall, this work reports that the binding modes of BH3 peptides are primarily dependent on its two terminal segments. The computational methods applied herein are also demonstrated to be of great assistance in the rational design of peptide inhibitors.