Helical Conformation In Solution Research Articles

A Rationally Designed Synthetic Antiviral Peptide Binder Targeting the Receptor-Binding Domain of SARS-CoV-2.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a novel coronavirus, is the causative agent responsible for the spread of the COVID19 pandemic across the globe. The global impact of the COVID19 pandemic, the successful approval of vaccines for controlling the pandemic, and the further resurgence of COVID19 necessitate the exploration and validation of alternative therapeutic avenues targeting SARS-CoV-2. The initial entry and further invasion by SARS-CoV-2 require strong protein-protein interactions (PPIs) between the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein and the human angiotensin-converting enzyme 2 (ACE2) receptors expressed on the cell surfaces of various tissues. In principle, disruption of the PPIs between the RBD of SARS-CoV-2 and the ACE2 receptor by designer peptides with optimized pharmacology appears to be an ideal choice for potentially preventing viral entry with minimal immunogenicity. In this context, the current study describes a short, synthetic designer peptide (codenamed SR16, ≤18 aa, molecular weight ≤2.5 kDa), which has a few noncoded amino acids, demonstrates a helical conformation in solution, and also engages the RBD of SARS-CoV-2 through a high-affinity interaction, as judged from a battery of biophysical studies. Further, the designer peptide demonstrates resistance to trypsin degradation, appears to be nontoxic to mammalian cells, and also does not induce hemolysis in freshly isolated human erythrocytes. In summary, SR16 appears to be an ideal peptide binder targeting the RBD of SARS-CoV-2, which has the potential for further optimization and development as an antiviral agent targeting SARS-CoV-2.

Hybrid Peptides Based on α-Aminoxy Acids as Antimicrobial and Anticancer Foldamers.

α-Aminoxy peptides represent an interesting group of peptidomimetics with high proteolytic stability and the ability to fold into specific, predictable secondary structures. Here, we present a series of hybrid peptides consisting of α-aminoxy acids and α-amino acids with cationic and aromatic, hydrophobic side chains in an alternating manner synthesized using an efficient protocol that combines solution- and solid-phase synthesis. 2D ROESY experiments with a representative hexamer suggested the presence of a 7/8 helical conformation in solution. Biological evaluation revealed a significant impact of the peptide chain length and the N-terminal cap on the antimicrobial and anticancer properties of this series of hybrid peptides. The Fmoc-capped peptide 6e displayed the most potent antimicrobial activity against a panel of Gram-negative and Gram-positive bacterial strains (e. g. against E. Coli: MIC=8 mg/L; S. aureus: MIC=4 mg/L).

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.

Rational design, conformational analysis and membrane-penetrating dynamics study of Bac2A-derived antimicrobial peptides against gram-positive clinical strains isolated from pyemia

The Bac2A (RLARIVVIRVAR−NH2) is a linearized counterpart of cationic cyclic peptide Bactenecin—one of the smallest naturally occurring antimicrobial peptides (AMPs), which, however, generally exhibits a low or moderate antibacterial potency against gram-positive bacteria. Here, it is found that the Bac2A and its linear derivates cannot spontaneously fold into a well-defined helical conformation in solution, thus impairing the peptide amphipathicity and antibacterial activity. Hydrocarbon stapling is rationally designed to constrain the helical conformation of these linear peptides. Atomistic dynamics simulations reveal that the membrane-penetrating course of linear and stapled peptides include four distinct phases, during which the stapled peptides can maintain in an ordered helical conformation, while linear peptides are structured from intrinsic disorder in water solution to partially helical state in membrane interior, indicating that lipid environment can help the linear peptide refolding into amphipathic helix, although the refolding process would incur a large configurational entropy penalty. The antibacterial activities of the most potent stapled peptide are determined as MIC = 7.6 and 16 µg/ml against two gram-positive Staphylococcus aureus clinical strains isolated from pyemia. The activity values are improved by 7.1-fold and 5-fold as compared to that of native Bac2A peptide with MIC = 54 and 80 µg/ml, respectively.

Synthesis and Conformational Analysis of Alternately N-Alkylated Aromatic Amide Oligomers.

Alternately N-alkylated aromatic amides such as 1-3 bearing various side chains were designed and synthesized as novel helical foldamers. The CD spectra of oligomers with chiral side chains showed a positive Cotton effect, which indicates that these oligomers take helical conformations in solution. The CD intensity gradually increased with increasing chain length, and pentamer 3d showed remarkably strong CD signals in chloroform. The absorption maxima of the UV spectra were increasingly red shifted with increasing chain length, in contrast to the case of poly( p- N-alkylbenzamide)s. Structure optimization of the oligomers based on the crystal structure of 1a as the monomer unit supported the formation of helical structure with a large cavity and also suggested intramolecular hydrogen bond formation between secondary amides. The results of calculation were consistent with the observed spectroscopic features.

Understanding the antimicrobial activity of water soluble γ-cyclodextrin/alamethicin complex.

In our previous investigation (Zhang et. al., J. Functional Foods 40 (2018) 700-706), we have proposed a method of complexation of alamethicin (ALM) with γ-cyclodextrin (γ-CD) to increase the solubility and demonstrated an enhancement in its antimicrobial activity against Listeria monocytogenes. In this study, transmission electron microscopy of L. monocytogenes treated with γ-CD/ALM complex indicated cell lysis due to pore formation. This was further corroborated by fluorescent dye leakage from DMPC/cholesterol liposomes when exposed to γ-CD/ALM complex for molar ratios of 1, 5 and 10. The extent of dye leakage increased with ALM-lipid ratio in the range of 0.00015 to 0.16. Dye leakage of γ-CD/ALM complex was found to be the highest for molar ratio of 5, consistent with our earlier results of antimicrobial activity of the complex against L. monocytogenes. All atom molecular dynamics (MD) simulation showed that γ-CD/ALM complex can effectively bind to the 3:1 POPE/POPG bilayer, a mimic of bacterial cell membrane. In addition, circular dichroism spectrum indicated that ALM in the complex has a helical conformation in solution as well as in the presence of liposome. Transmembrane MD simulation of six γ-CD/ALM complex aggregates in α helical conformation showed water channel with barrel stave like pore structure.

Lycium europaeum Linn as a source of polysaccharide with in vitro antioxidant activities and in vivo anti-inflammatory and hepato-nephroprotective potentials

Ethnopharmacological relevanceLycium europaeum Linn. is widely used to treat the burning of the skin and well-known as a medicinal plant having various biological activities. Aims of the studyThe purpose of the present study is to characterize the polysaccharide from L. europaeum L. leaves (LEP) and to explore its antioxidant, anti-inflammatory and hepato-nephroprotective properties. Materials and methodsThe structural and functional characteristics of LEP were investigated using X-ray diffraction techniques (XRD), Scanning Electron Microscopy (SEM), and FT-IR Spectroscopy. The antioxidant activity was evaluated using 2,2-diphenyl-1-picrylhydrazyl and hydrogen peroxide scavenging assays. Hepato-renal effects were studied using CCl4 and cisplatin-induced liver and kidney injuries in mice, respectively. Anti-inflammatory activity was assessed on carrageenan-induced paw edema. ResultsThe LEP showed an interesting water-holding capacity and effective foaming and emulsifying properties. XRD analysis suggested that LEP form a semi-crystalline polymer with an amorphous structure. FT-IR profile showed the presence of pyranose ring in LEP. SEM and helix-coil transition analyses indicated that LEP had a lamellar structure with angular edges and didn't present a triple helical conformation in solution. In vitro, LEP indicated significant concentration-dependent antioxidant activity. In vivo, LEP treatment significantly reduced the effects of CCl4 intoxication on serum liver biomarkers (AST, ALT, LDH, and GGT) and the effect of cisplatin on serum renal biomarkers (urea, blood urea nitrogen, creatinine, and uric acid). Meanwhile, LEP diminishes significantly the effect of CCl4 and cisplatin on the level of lipid peroxidation in liver and kidney tissues, respectively. Additionally, the normal histological structure of liver and kidney was restored after treatment with the polysaccharide. LEP possessed a significant anti-inflammatory activity on acute inflammation induced by carrageenan in mice. ConclusionOverall, the findings of this study support the traditional use of L. europaeum L. This plant may also be used as a good agent for protection against inflammatory diseases and hepato-renal injuries in patients with cancer.

Biologically Active Branched Polysaccharide Mimetics: Synthesis via Ring-Opening Polymerization of a Maltose-Based β-Lactam.

Stereoregular poly-amido-saccharides bearing α-glucopyranose branches (Mal-PASs) are synthesized by anionic ring-opening polymerization of a maltose-based β-lactam monomer followed by debenzylation. The polymerization affords high molecular weight polymers (up to 31500 g/mol) with narrow dispersities (Đ < 1.1). Deprotected Mal-PASs are highly soluble in water and adopt a left-handed helical conformation in solution. Turbidimetric assay shows that Mal-PASs are multivalent ligands to lectin Concanavalin A.

Induction of high titred, non-neutralising antibodies by self-adjuvanting peptide epitopes derived from the respiratory syncytial virus fusion protein

Respiratory syncytial virus (RSV) causes severe lower respiratory tract illness in infants and young children. The significant morbidity and mortality rates associated with RSV infection make an effective RSV vaccine development a priority. Two neutralising antibody binding sites, Ø and II, located on the pre-fusion RSV F glycoprotein are prime candidates for epitope-focused vaccine design. We report on a vaccine strategy that utilises a lipid core peptide (LCP) delivery system with self-adjuvanting properties in conjunction with either the antigenic site Ø or II (B cell epitopes) along with PADRE as a T helper cell epitope. These LCP constructs adopted the desired helical conformation in solution and were recognised by their cognate antibodies D25 and Motavizumab, specific for site Ø and II on RSV F protein, respectively. The LCP constructs were capable of eliciting higher levels of antigen specific antibodies than those induced by antigens administered with complete Freund’s adjuvant, demonstrating the potent adjuvanting properties of LCP delivery. However, the antibodies induced failed to recognise native F protein or neutralise virus infectivity. These results provide a note of caution in assuming that peptide vaccines, successfully designed to structurally mimic minimal linear B cell epitopes, will necessarily elicit the desired immune response.

Helical Folding of π-Conjugated Polymers Bearing Glucose-Linked Biphenyl Units in the Main Chain: Application to Circularly Polarized Luminescence Materials

Abstract We have synthesized a series of chiral π-conjugated polymers bearing glucose-linked biphenyl units in the main chain through copolymerizations of a key diethynyl compound (GLB-1), containing the structural element of naturally-occurring ellagitannin, with various diiodoaryls. The circular dichroism and circularly polarized luminescence properties of the obtained polymers were investigated in solution and the solid state. Based on a comparative study using the corresponding unimer model compounds, poly-3 and poly-4 bearing (oligo)thiophene-type comonomer units are considered to be able to fold into an excess one-handed helical conformation in solution and the solid state under an appropriate exterior environment. In contrast, analogous polymers bearing p- and m-phenylene residues as comonomer units did not form regular higher-ordered structures. We also observed that poly-3 and poly-4 efficiently emitted left-handed circularly polarized light with a corresponding fluorescence region arising from their helical chirality, and possessed dissymmetry factors of approximately 1.0 × 10−2 for the film state.

Molecular Environment Modulates Conformational Differences between Crystal and Solution States of Human β-Defensin 2.

Human β-defensin 2 is a cysteine-rich antimicrobial peptide. In the crystal state, the N-terminal segment (residues 1-11) exhibits a helical conformation. However, a truncated form, with four amino acids removed from the N-terminus, adopts nonhelical conformations in solution, as shown by NMR. To explore the molecular origins of these different conformations, we performed Hamiltonian replica exchange molecular dynamics simulations of the peptide in solution and in the crystal state. It is found that backbone hydration and specific protein-protein interactions are key parameters that determine the peptide conformation. The helical conformation in the crystal state mainly arises from reduced hydration as well as a salt bridge between the peptide and a symmetry-related neighboring monomer in the crystal. When the extent of hydration is reduced and the salt bridge is reintroduced artificially, the peptide is successfully folded back to the helical conformation in solution. The findings not only shed light on the development of accurate force field parameters for protein molecules but also provide practical guidance in the design of functional proteins and peptides.

12/10-Helical β-Peptide with Dynamic Folding Propensity: Coexistence of Right- and Left-Handed Helices in an Enantiomeric Foldamer.

We present the first examples of atomic-resolution crystal data for the β-peptide 12/10-helix from oligomers of cis-2-aminocyclohexane carboxylic acid (cis-ACHC) with alternating chirality. The local conformations of two enantiomeric cis-ACHC dimer units suggested that a chiral β-peptide may adopt both right-handed and left-handed helical conformations in solution. To probe the conformational behavior of 12/10-helical β-peptides, the two reference helices with a single handedness were synthesized with a more rigidified cis-ACHC derivative. Comparison with these reference helices at low temperature revealed that a chiral cis-ACHC oligomer with alternating chirality indeed displays 12/10-helical conformations with both handedness that equilibrate rapidly in solution. This is a very rare example of chiral oligomers with helix inversion ability. The 12/10-helical backbone should be a valuable addition to potential scaffolds for applications involving helices with dynamic folding propensity.

Conformational Flexibility Enables Function of a BECN1 Region Essential for Starvation‐Mediated Autophagy

BECN1 is essential for autophagy, a critical eukaryotic cellular homeostasis pathway. Here we delineate a highly conserved BECN1 domain (residues 141–171), located between the previously characterized BH3 and coiled‐coil domains, and elucidate its structure and role in autophagy. The 2.0 Å Sulfur‐Single Wavelength Anomalous Diffraction crystal structure of this domain demonstrates that its N‐terminal half is unstructured, while its C‐terminal half is helical, hence we name it the Flexible Helical Domain (FHD). Circular dichroism, Double‐Electron‐Electron Resonance‐Electron Paramagnetic Resonance and Small Angle X‐ray Scattering (SAXS) analyses confirm that this domain is partially disordered even in the context of adjacent BECN1 domains. Molecular dynamic simulations fitted to SAXS data indicate that the FHD transiently samples more helical conformations in solution. Strikingly, the FHD is predicted to fold upon binding, and we find it becomes more helical in the presence 2,2,2‐trifluoroethanol, indicating it may undergo a disorder‐to‐helix conformational change upon binding to a partner. A completely helical conformation would place highly conserved FHD residues on the same face of the helix, suggesting that these residues may be involved in binding. Lastly, cellular studies show that highly conserved FHD residues are essential for the starvation‐induced up‐regulation of autophagy, the first BECN1 region shown to be critical for starvation‐induced autophagy rather than basal autophagy. Thus, our combined data suggest that the FHD likely undergoes a disorder‐to‐helix transition upon binding to appropriate partners, and conserved residues critical for this interaction are essential for starvation‐induced up‐regulation of autophagy.Support or Funding InformationThis work was supported by National Institutes of Health grants RO3 NS090939 (S.S.) and R15 GM113227 (C.C.); a National Science Foundation grant MCB‐1413525 (S.S.); North Dakota EPSCoR doctoral dissertation award for Y.M. (S.S.). AR was supported by a ORNL Laboratory Director's Research and Development SEED proposal 7278.

Helix-sense-selective radical polymerization of (S)-6-acryloyl-2,2′-bisalkoxy-1,1′-binaphthyl

A series of (S)-6-acryloyl-2,2′-bisalkoxy-1,1′-binaphthyl 1 were radically polymerized to obtain polymers. The optical rotation and the circular dichroism (CD) signal intensity of the radically polymerized poly-1 were close to that of the anionically polymerized poly-1 indicating that the radically polymerized poly-1 should adopt stable preferred-handed helical conformation in solution just as the anionically polymerized poly-1.

Optically Active Helical Poly[(S)‐3‐vinyl‐2,2′‐dihydroxy‐1,1′‐binaphthyl]: New Ligand for Asymmetric Addition of Diethylzinc to Aryl Aldehyde

AbstractPoly[(S)‐3‐vinyl‐2,2′‐dihydroxy‐1,1′‐binaphthyl] (L*) was obtained by taking off the protecting groups of poly[(S)‐3‐vinyl‐2,2′‐bis(methoxymethoxy)‐1,1′‐binaphthyl] (poly‐1). L* was proved to keep a stable helical conformation in solution. The application of helical L* in the asymmetric addition of diethylzinc to aldehydes has been studied. The catalytic system employing 10 mol% of L* and 150 mol% of Ti(OiPr)4 was found to promote the addition of diethylzinc to a wide range of aromatic aldehydes, giving up to 99% enantiomeric excess (ee) and up to 93% yield of the corresponding secondary alcohol at 0°C. The chiral polymer can be easily recovered and reused without loss of catalytic activity as well as enantioselectivity.

Detection of Listeria monocytogenes with short peptide fragments from class IIa bacteriocins as recognition elements.

We employed a direct peptide-bacteria binding assay to screen peptide fragments for high and specific binding to Listeria monocytogenes. Peptides were screened from a peptide array library synthesized on cellulose membrane. Twenty four peptide fragments (each a 14-mer) were derived from three potent anti-listerial peptides, Leucocin A, Pediocin PA1, and Curvacin A, that belong to class IIa bacteriocins. Fragment Leu10 (GEAFSAGVHRLANG), derived from the C-terminal region of Leucocin A, displayed the highest binding among all of the library fragments toward several pathogenic Gram-positive bacteria, including L. monocytogenes, Enterococcus faecalis, and Staphylococcus aureus. The specific binding of Leu10 to L. monocytogenes was further validated using microcantilever (MCL) experiments. Microcantilevers coated with gold were functionalized with peptides by chemical conjugation using a cysteamine linker to yield a peptide density of ∼4.8×10(-3) μmol/cm2 for different peptide fragments. Leu10 (14-mer) functionalized MCL was able to detect Listeria with same sensitivity as that of Leucocin A (37-mer) functionalized MCL, validating the use of short peptide fragments in bacterial detection platforms. Fragment Leu10 folded into a helical conformation in solution, like that of native Leucocin A, suggesting that both Leu10 and Leucocin A may employ a similar mechanism for binding target bacteria. The results show that peptide-conjugated microcantilevers can function as highly sensitive platforms for Listeria detection and hold potential to be developed as biosensors for pathogenic bacteria.

Exploring the consequences of a representative "disallowed" conformation of Aib on a 3₁₀-helical fold.

The structural effects of a representative "disallowed" conformation of Aib on the 3(10)-helical fold of an octapeptidomimetic are explored. The 1D ((1)H, (13)C) & 2D NMR, FT-IR and CD data reveal that the octapeptide 1, adopts a 3(10)-helical conformation in solution, as it does in its crystal structure. The C-terminal methyl carboxylate (CO2Me) of 1 was modified into an 1,3-oxazine (Oxa) functional group in the peptidomimetic 2. This modification results in the stabilization of the backbone of the C-terminal Aib (Aib*-Oxa) of 2, in a conformation (ϕ, ψ = 180, 0) that is natively disallowed to Aib. Consequent to the presence of this natively disallowed conformation, the 3(10)-helical fold is not disrupted in the body of the peptidomimetic 2. But the structural distortions that do occur in 2 are primarily in residues in the immediate vicinity of the natively disallowed conformation, rather than in the whole peptide body. Non-native electronic effects resulting from modifications in backbone functional groups can be at the origin of stabilizing residues in natively disallowed conformations.

Optically active helical polyethers from chiral terminal epoxides with a diarylamino group

Six optically active epoxy propanes with a diarylamino group at the 3-position carbon were synthesized and polymerized. Poly((S)-3-diphenylaminopropene oxide) ([Formula: see text] = –634°) was obtained from the polymerization of (S)-3-diphenylaminopropene oxide ([Formula: see text] = +75°). By investigating chiroptical properties of the obtained polyethers derived from optically active epoxy propanes, we found that if the diarylamino group were rigid enough, the obtained polyethers from chiral terminal epoxides with a diarylamino group could maintain a one-handed helical conformation in solution.

Synthesis, characterization, polymerization of optically active terminal epoxides and episulfides bearing bulky groups

A simple and efficient protocol was developed for the syntheses of optically active epoxides(2a–2d) via the reactions of epichlorohydrin with organometallic reagents with steric hindrance at low temperature. On this basis, episulfides(3a–3d) were obtained with high e.e. values by the reaction of the corresponding epoxides with NH4SCN. The configuration of thiiranes was opposite to that of the corresponding epoxides. Then, epoxides 2a–2d were anionically polymerized and the poly-R-2a(or poly-S-2a) was proved to keep a stable one-handed helical conformation in solution. The oligomers[degree of polymerization(DP)=2–5] were obtained when the thiiranes(3a, 3b and 3c) were used to polymerize.

Bispidine as a helix inducing scaffold: examples of helically folded linear peptides

We designed and synthesized bispidine-anchored peptides and showed that these peptides as small as (containing four chiral α-amino acid residues) adopt a right handed helical conformation. Bispidine anchored linear peptide adopts a helical conformation in solution and in the solid state.