Conformation In Aqueous Solution Research Articles

Favorable Symmetric Structures of Radiopharmaceutically Important Neutral Cyclen-Based Ligands

Cyclen-based ligands are prominent tools for transferring radioisotopes through the human body. A crucial criterion is the stability of their complexes, which is partly determined by the stabilization of the free ligand in solution. For the assessment of the later property, the favored conformation(s) in the solution must be known. In the present study, the conformational space of four neutral cyclen-based ligands was elucidated by a multi-step procedure: the survey of the conformational space using molecular mechanics (MM) was followed by Density Functional Theory (DFT) calculations on the low-energy conformers and evaluation of the solvent effects. The results revealed several low-energy conformers in aqueous solution. In terms of electronic energies, a significant preference of symmetric structures (C4 or C2—similar to the ligand arrangements in their metal complexes) was obtained. The thermal contributions to the Gibbs free energy (mainly the vibrational ones) tend to decrease this preference by several kJ/mol against non-symmetric structures. Nonetheless, the advantage of compact symmetric structures was confirmed in all the four studied cases.

Structural characteristics, immune-activating mechanisms in vitro, and immunomodulatory effects in vivo of the exopolysaccharide EPS53 from Streptococcus thermophilus XJ53

Our previous investigations have successfully identified the repeating structural units of EPS53, an exopolysaccharide derived from Streptococcus thermophilus XJ53 fermented milk, and substantiated its potential immunomodulatory properties. The present study further elucidated the structural characteristics of EPS53 and investigated the underlying mechanisms governing its in vitro immunoreactivity as well as its in vivo immunoreactivity. The results obtained from multi-detector high performance gel filtration chromatography revealed that EPS53 adopted a rigid rod conformation in aqueous solution, with the weight-average molecular weight of 1464 kDa, the number-average molecular weight of 694 kDa, and the polydispersity index of 2.11. Congo red experiment confirmed the absence of a triple helix conformation. Scanning electron microscopy showed that EPS53 displayed a three-dimensional fibrous structure covered with flakes. The in vitro findings indicated that EPS53 enhanced phagocytosis ability, reactive oxygen species (ROS) production, and cytokine levels of macrophages via the TLR4-mediated NF-κB/MAPK signaling pathways as confirmed by immunofluorescence staining experiments, inhibition blocking experiments, and Western blot assay. Additionally, the in vivo experiments demonstrated that EPS53 significantly increased macrophage and neutrophil number while enhancing NO and ROS levels in zebrafish larvae; thus, providing further evidence for the immunomodulatory efficacy of EPS53.

Does Inter-Residue Hydrogen Bonding in β-(1→4)-Linked Disaccharides Influence Linkage Conformation in Aqueous Solution?

Two disaccharides, methyl β-d-galactopyranosyl-(1→4)-α-d-glucopyranoside (1) and methyl β-d-galactopyranosyl-(1→4)-3-deoxy-α-d-ribo-hexopyranoside (3), were prepared with selective 13C-enrichment to allow measurement of six trans-O-glycosidic J-couplings (2JCOC, 3JCOCH, and 3JCOCC) in each compound. Density functional theory (DFT) was used to parameterize Karplus-like equations that relate these J-couplings to either ϕ or ψ. MA'AT analysis was applied to both linkages to determine mean values of ϕ and ψ in each disaccharide and their associated circular standard deviations (CSDs). Results show that deoxygenation at C3 of 1 has little effect on both the mean values and librational motions of the linkage torsion angles. This finding implies that, if inter-residue hydrogen bonding between O3H and O5' of 1 is present in aqueous solution and persistent, it plays little if any role in dictating preferred linkage conformation. Hydrogen bonding may lower the energy of the preferred linkage geometry but does not determine it to any appreciable extent. Aqueous 1-μs MD simulation supports this conclusion and also indicates greater conformational flexibility in deoxydisaccharide 3 in terms of sampling several, conformationally distinct, higher-energy conformers in solution. The populations of these latter conformers are low (3-14%) and could not be validated by MA'AT analysis. If the MD model is correct, however, C3 deoxygenation does enable conformational sampling over a wider range of ϕ/ψ values, but linkage conformation in the predominant conformer is essentially identical in both 1 and 3.

Conformational control of antimicrobial peptide amphiphilicity: consequences for boosting membrane interactions and antimicrobial effects of photocatalytic TiO2 nanoparticles.

This study reports on the effects of conformationally controlled amphiphilicity of antimicrobial peptides (AMPs) on their ability to coat TiO2 nanoparticles (NPs) and boost the photocatalytic antimicrobial effects of such NPs. For this, TiO2 NPs were combined with AMP EFK17 (EFKRIVQRIKDFLRNLV), displaying a disordered conformation in aqueous solution but helix formation on interaction with bacterial membranes. The membrane-bound helix is amphiphilic, with all polar and charged amino acid residues located at one side and all non-polar and hydrophobic residues on the other. In contrast, the d-enantiomer variant EFK17-d (E(dF)KR(dI)VQR(dI)KD(dF)LRNLV) is unable to form the amphiphilic helix on bacterial membrane interaction, whereas the W-residues in EFK17-W (EWKRWVQRWKDFLRNLV) boost hydrophobic interactions of the amphiphilic helix. Circular dichroism results showed the effects displayed for the free peptide, to also be present for peptide-coated TiO2 NPs, causing peptide binding to decrease in the order EFK17-W > EFK17 > EFK17-d. Notably, the formation of reactive oxygen species (ROS) by the TiO2 NPs was essentially unaffected by the presence of peptide coating, for all the peptides investigated, and the coatings stabilized over hours of UV exposure. Photocatalytic membrane degradation from TiO2 NPs coated with EFK17-W and EFK17 was promoted for bacteria-like model bilayers containing anionic phosphatidylglycerol but suppressed in mammalian-like bilayers formed by zwitterionic phosphatidylcholine and cholesterol. Structural aspects of these effects were further investigated by neutron reflectometry with clear variations observed between the bacteria- and mammalian-like model bilayers for the three peptides. Mirroring these results in bacteria-like model membranes, combining TiO2 NPs with EFK17-W and EFK17, but not with non-adsorbing EFK17-d, resulted in boosted antimicrobial effects of the resulting cationic composite NPs already in darkness, effects enhanced further on UV illumination.

Discovery of structural and functional transition sites for membrane-penetrating activity of sheep myeloid antimicrobial peptide-18

Cathelicidin antimicrobial peptides have an extended and/or unstructured conformation in aqueous solutions but fold into ordered conformations, such as the α-helical structure, when interacting with cellular membranes. These structural transitions can be directly correlated to their antimicrobial activity and its underlying mechanisms. SMAP-18, the N-terminal segment (residues 1–18) of sheep cathelicidin (SMAP-29), is known to kill microorganisms by translocating across membranes and interacting with their nucleic acids. The amino acid sequence of SMAP-18 contains three Gly residues (at positions 2, 7, and 13) that significantly affect the flexibility of its peptide structure. This study investigated the role of Gly residues in the structure, membrane interaction, membrane translocation, and antimicrobial mechanisms of SMAP-18. Five analogs were designed and synthesized through Gly → Ala substitution (i.e., G2A, G7A, G13A, G7,13A, and G2,7,13A); these substitutions altered the helical content of SMAP-18 peptides. We found that G7,13A and G2,7,13A changed their mode of action, with circular dichroism and nuclear magnetic resonance studies revealing that these analogs changed the structure of SMAP-18 from a random coil to an α-helical structure. The results of this experiment suggest that the Gly residues at positions 7 and 13 in SMAP-18 are the structural and functional determinants that control its three-dimensional structure, strain-specific activity, and antimicrobial mechanism of action. These results provide valuable information for the design of novel peptide-based antibiotics.

Fabrication of salt-tolerant chitosan-based polyelectrolyte flocculant through enhancing H-bond hydration effect for treating and recycling of highly saline dyeing wastewater

The textile industry is required to execute water conservation and contamination management techniques owing to increasingly serious water scarcity and pollution. Herein, a novel salt-tolerant chitosan-based flocculant (CT-CTA-BGE3) with high charge density was synthesized through chitosan (CT) etherification with 2,3-epoxypropyl trimethylammonium chloride (CTA) and butyl glycidyl ether (BGE). The H-bond hydration effect between BGE segments and water molecules weakens the salt-induced polyelectrolyte effect, allowing CT-CTA-BGE3 to chains keep an extended conformation in high-salty aqueous solution and thus endowing excellent salt-tolerant properties. Meanwhile, the incorporation of CTA segments offers a high positive charge on the flocculant surface. Thus, the beneficial effect of satisfying color removal was obtained by CT-CTA-BGE3 in a highly saline system because it would render the more exposed active sites with strong positive charges along stretched polymer chains. The outstanding flocculability of CT-CTA-BGE3 (3054 mg/g) for the reactive brilliant red 2 (a widely commercial grade dye) could be obtained within 5 min (pH = 6). As expected, because the CT-CTA-BGE3 impurities can be reduced in reclaimed water via the pH-mediated hydrophilicity-hydrophobicity switch of polymer, the obtained clarified saline wastewater after flocculation can be recycled in a new dyeing bath without sacrificing fabric quality (ΔE*less than1), and over 88 % of NaCl can be regenerated for each cycle. This work develops an alternative strategy for the fabrication of salt-tolerant flocculants through enhancing H-bond hydration, revealing the promising application prospects in treating and recycling highly saline textile wastewater.

Locust Bean Gum, a Vegetable Hydrocolloid with Industrial and Biopharmaceutical Applications

Locust bean gum (LBG), a vegetable galactomannan extracted from carob tree seeds, is extensively used in the food industry as a thickening agent (E410). Its molecular conformation in aqueous solutions determines its solubility and rheological performance. LBG is an interesting polysaccharide also because of its synergistic behavior with other biopolymers (xanthan gum, carrageenan, etc.). In addition, this hydrocolloid is easily modified by derivatization or crosslinking. These LBG-related products, besides their applications in the food industry, can be used as encapsulation and drug delivery devices, packaging materials, batteries, and catalyst supports, among other biopharmaceutical and industrial uses. As the new derivatized or crosslinked polymers based on LBG are mainly biodegradable and non-toxic, the use of this polysaccharide (by itself or combined with other biopolymers) will contribute to generating greener products, considering the origin of raw materials used, the modification procedures selected and the final destination of the products.

Optimization of ultrasound-assisted aqueous two-phase extraction of polysaccharides from seabuckthorn fruits using response methodology, physicochemical characterization and bioactivities.

Seabuckthorn fruits contains many active subtances, among them, the seabuckthorn polysaccharide is one of the main active ingredients, and exhibits diverse bioactivities. The extraction of polysaccharides from seabuckthorn fruits is the most important step for their wide applications. Ultrasound-assisted aqueous two-phase extraction (UA-ATPE) is a promising green method for extracting polysaccharides. Additionally, physicochemical characterization and antioxidant activities can evaluate the potential functions and applications in the food and medicine industries. Based on the single-factor experiments, 20.70% (w/w) ammonium sulfate ((NH4 )2 SO4 ) and 27.56% (w/w) ethanol were determined as the suitable composition for aqueous two-phase. The optimum conditions of UA-ATPE obtained by response surface methodology were as follows: ultrasonic power (390 W), extraction time (41 min), liquid-to-material ratio (72: 1mL/g), and the total yield of the polysaccharides reached 34.14 ± 0.10%, The molecular weights of the purified upper-phase seabuckthorn polysaccharide (PUSP) and the purified lower-phase seabuckthorn polysaccharide (PLSP) were 65 525 and 26 776 Da, respectively. PUSP and PLSP contained the same six monosaccharides (galacturonic acid, rhamnose, xylose, mannose, glucose and galactose), but with different molar ratios. Furthermore, PUSP and PLSP displayed certain viscoelastic property, had no triple helical structure, possessed different thermal stability, surface morphology and conformation in aqueous solution. PUSP and PLSP displayed strong antioxidant properties by the assays of scavenging ability of ABTS+ ·, the protection of DNA damage and erythrocyte hemolysis. UA-ATPE significantly increased the yield of seabuckthorn polysaccharides. PUSP and PLSP were different in many aspects, such as molar ratio, surface shape and antioxidant activities. Seabuckthornpolysaccharides possess great potential in medicine and functional foods. © 2022 Society of Chemical Industry.

Membrane Association Allosterically Regulates Phospholipase A2 Enzymes and Their Specificity.

Water-soluble proteins as well as membrane-bound proteins associate with membrane surfaces and bind specific lipid molecules in specific sites on the protein. Membrane surfaces include the traditional bilayer membranes of cells and subcellular organelles formed by phospholipids. Monolayer membranes include the outer monolayer phospholipid surface of intracellular lipid droplets of triglycerides and various lipoproteins including HDL, LDL, VLDL, and chylomicrons. These lipoproteins circulate in our blood and lymph systems and contain triglycerides, cholesterol, cholesterol esters, and proteins in their interior, and these are sometimes interspersed on their surfaces. Similar lipid-water interfaces also occur in mixed micelles of phospholipids and bile acids in our digestive system, which may also include internalized triglycerides and cholesterol esters. Diacyl phospholipids constitute the defining molecules of biological membranes. Phospholipase A1 (PLA1) hydrolyzes phospholipid acyl chains at the sn-1 position of membrane phospholipids, phospholipase A2 (PLA2) hydrolyzes acyl chains at the sn-2 position, phospholipase C (PLC) hydrolyzes the glycerol-phosphodiester bond, and phospholipase D (PLD) hydrolyzes the polar group-phosphodiester bond. Of the phospholipases, the PLA2s have been the most well studied at the mechanistic level. The PLA2 superfamily consists of 16 groups and numerous subgroups, and each is generally described as one of 6 types. The most well studied of the PLA2s include extensive genetic and mutational studies, complete lipidomics specificity characterization, and crystallographic structures. This Account will focus principally on results from deuterium exchange mass spectrometric (DXMS) studies of PLA2 interactions with membranes and extensive molecular dynamics (MD) simulations of their interactions with membranes and specific phospholipids bound in their catalytic and allosteric sites. These enzymes either are membrane-bound or are water-soluble and associate with membranes before extracting their phospholipid substrate molecule into their active site to carry out their enzymatic hydrolytic reaction. We present evidence that when a PLA2 associates with a membrane, the membrane association can result in a conformational change in the enzyme whereby the membrane association with an allosteric site on the enzyme stabilizes the enzyme in an active conformation on the membrane. We sometimes refer to this transition from a "closed" conformation in aqueous solution to an "open" conformation when associated with a membrane. The enzyme can then extract a single phospholipid substrate into its active site, and catalysis occurs. We have also employed DXMS and MD simulations to characterize how PLA2s interact with specific inhibitors that could lead to potential therapeutics. The PLA2s constitute a paradigm for how membranes interact allosterically with proteins, causing conformational changes and activation of the proteins to enable them to extract and bind a specific phospholipid from a membrane for catalysis, which is probably generalizable to intracellular and extracellular transport and phospholipid exchange processes as well as other specific biological functions. We will focus on the four main types of PLA2, namely, the secreted (sPLA2), cytosolic (cPLA2), calcium-independent (iPLA2), and lipoprotein-associated PLA2 (Lp-PLA2) also known as platelet-activating factor acetyl hydrolase (PAF-AH). Studies on a well-studied specific example of each of the four major types of the PLA2 superfamily demonstrate clearly that protein subsites can show precise specificity for one of the phospholipid hydrophobic acyl chains, often the one at the sn-2 position, including exquisite sensitivity to the number and position of double bonds.

Probing the effect of NEK7 and cofactor interactions on dynamics of NLRP3 monomer using molecular simulation.

The NLRP3 inflammasome is a cytoplasmic complex that regulates the activation of inflammatory cytokines and, given its implication in a range of diseases, is an important therapeutic target. The cofactor ATP and the centrosomal kinase NEK7 are important for NLRP3 activation. Here we have constructed and simulated computational models of full‐length monomeric NLRP3 to shed light on the importance of NEK7 and cofactor interactions for its conformation and dynamics in aqueous solution. We find that molecular dynamics simulation reproduces well the features of the recently published cryo‐EM structure of the ADP‐bound NLRP3–NEK7 complex; on the removal of NEK7, the NLRP3 molecule adopts a more compact closed form during simulations. Replacement of ADP by ATP promotes a rearrangement of hydrogen‐bonding interactions, domain interfaces, and a degree of opening of the NLRP3 conformation. We also examine the dynamics of an acidic loop of the LRR domain of NLRP3, which samples in a region observed in the NEK7‐bound cryo‐EM structure but not in an oligomeric form of inactive NLRP3. During the molecular dynamics simulations of NLRP3, we find some plasticity in its topology that suggests access routes for ATP to the cofactor pocket not immediately evident from the existing NEK7‐bound cryo‐EM structure. These computed dynamical trajectories of NLRP3 provide insight into coordinates of deformation that may be key for cofactor binding and inflammasome activation.

Characterization and macrophages immunomodulatory activity of two water-soluble polysaccharides from Abrus cantoniensis.

The study aims to elucidate the physicochemical properties and immunomodulatory activity of two polysaccharides (ACPt0 and ACPt2) from Abrus cantoniensis. Results revealed that ACPt0 with a molecular weight of 26.0 kDa, was mainly composed of glucose (83.1%) and galactose (6.1%), and that ACPt2 with a molecular weight of 145.6/8.9 kDa, consisted of galactose (25.6%), galacturonic acid (22.2%), arabinos (16.6%) and galactose (11.0%) respectively. AFM and Congo red experiments suggested that ACPt0 and ACPt2 might be spherical particles with triple-helix conformation in aqueous solution. ACPt0 and ACPt2 exhibited immunomodulatory activity by promoting the proliferation, augmenting pinocytic and phagocytic capacities, releasing immunoactive molecules such as ROS, NO, iNOS, TNF-α, IL-6 and IL-1β, upregulation of the mRNA levels of corresponding cytokines in macrophages. Moreover, ACPt0 and ACPt2 were recognized by toll-like receptor 4 (TLR4) and exerted immunomodulatory effects via activating Myeloid differentiation factor 88 (MyD88), mitogen-activated protein kinases (MAPKs) and serine/threonine kinase (Akt) signaling pathways in macrophages. Notably, ACPt2 had higher immunomodulatory activity than ACPt0. Based on the present findings, ACPt0 and ACPt2 could be explored as an active component of immunomodulators in the food and pharmaceutical fields.

Effects of steam on polysaccharides from Polygonatum cyrtonema based on saccharide mapping analysis and pharmacological activity assays

BackgroundPolygonatum cyrtonema, one of origins of Polygonata Rhizoma (HuangJing in Chinese), is traditionally steamed repeatedly before being used as herbal medicine in China. However, there has no standard for steaming of HuangJing. Therefore, a comprehensive study for effects of steam on polysaccharides from Polygonatum cyrtonema based on saccharide mapping, a powerful method developed for polysaccharides analysis, and pharmacological activity are still necessary, which is helpful to explore the effect of steam on the physiochemical and biological activities of its polysaccharides and develop steaming standard of Polygonatum cyrtonema.MethodsTo explore the effect of steam on physiochemical and biological activities of P. cyrtonema polysaccharides (PCP), six polysaccharides named PCP0, PCP1, PCP2, PCP3, PCP4 and PCP5 were extracted from the herb consecutively steamed for 0–5 times, respectively. Their molecular weight distribution, monosaccharide composition and PACE fingerprints were investigated through HPSEC-MALLS-RID, HPAEC-PAD and saccharide mapping based on polysaccharides analysis by using carbohydrate gel electrophoresis (PACE) and HPTLC, respectively. In addition, their antioxidant ability and immunostimulatory activities on RAW 264.7 cells in terms of NO production and phagocytosis were compared.ResultsResults suggested that molecular weights could be changed during steam, which increased by first steaming and then decreased with further steaming though all polysaccharides’ molecular weights were 105-107 Da. They all showed irregularly spherical conformation in aqueous solution based on AFM imaging. Their monosaccharide composition and PACE fingerprints were significantly different after steaming, i.e., galactose increased while glucose and mannose decreased, and β-1,4-Galp appeared while β-1,4-Manp increased, after steaming. Steamed PCP significantly increased scavenging activity against ABTS radicals, while PCP0 had the best immunostimulatory effect on RAW 264.7 in terms of NO production and phagocytosis.ConclusionsIn summary, steam significantly affected the chemical composition and bioactivities of polysaccharides from P. cyrtonema. Considering the balance beneficial effects of steaming on antioxidant and immunopotentiation activities of PCP, 2 times of continuous steam is the optimal choice under the given conditions.

Antimicrobial Polymers of Linear and Bottlebrush Architecture: Probing the Membrane Interaction and Physicochemical Properties

Polymeric antimicrobial peptide mimics are a promising alternative for the future management of the daunting problems associated with antimicrobial resistance. However, the development of successful antimicrobial polymers (APs) requires careful control of factors such as amphiphilic balance, molecular weight, dispersity, sequence, and architecture. While most of the earlier developed APs focus on random linear copolymers, the development of APs with advanced architectures proves to be more potent. It is recently developed multivalent bottlebrush APs with improved antibacterial and hemocompatibility profiles, outperforming their linear counterparts. Understanding the rationale behind the outstanding biological activity of these newly developed antimicrobials is vital to further improving their performance. This work investigates the physicochemical properties governing the differences in activity between linear and bottlebrush architectures using various spectroscopic and microscopic techniques. Linear copolymers are more solvated, thermo-responsive, and possess facial amphiphilicity resulting in random aggregations when interacting with liposomes mimicking Escheria coli membranes. The bottlebrush copolymers adopt a more stable secondary conformation in aqueous solution in comparison to linear copolymers, conferring rapid and more specific binding mechanism to membranes. The advantageous physicochemical properties of the bottlebrush topology seem to be a determinant factor in the activity of these promising APs.

Research on Extraction, Structure Characterization and Immunostimulatory Activity of Cell Wall Polysaccharides from Sparassis latifolia.

The cell wall polysaccharides were extracted from Sparassis latifolia fruit bodies by acid–alkali and superfine-grinding assisted methods, and the chemical characterization and in vitro immunity activities of these polysaccharide fractions were studied and compared. Results showed that superfine-grinding assisted extraction exhibited the highest yield of polysaccharides (SP, 20.80%) and low β-glucan content (19.35%) compared with alkaline extracts. The results revealed that the 20% ethanol precipitated fraction (20E) from SP was mainly composed of β-(1→3)-glucan and α-(1→4)-glucan. With the increase of ethanol precipitation, the fractions (30E, 40E, 50E) were identified as α-(1→4)-glucan with different molecular weights and conformations. Cell wall polysaccharides extracted through NaOH (NSP) and KOH (KSP) extraction had similar yields with 8.90% and 8.83%, respectively. Structural analysis indicated that the purified fraction from KSP (KSP-30E) was a β-(1→3)-glucan backbone branched with β-(1→6)-Glcp, while the purified fraction from NSP (NSP-30E) mainly contained β-(1→3)-glucan with a small number of α-linked-Glcp. The two fractions both exhibited rigid chain conformation in aqueous solutions. All polysaccharide fractions exerted the activity of activating Dectin-1 receptor in vitro, and the KSP-30E mainly identified as β-(1→3)-glucan with the terminal group via 1→6-linkage attached at every third residue exhibited a stronger enhancing effect than other fractions. Results suggested that KOH extraction could be efficient for the preparation of bioactive β-(1→3, 1→6)-glucan as a food ingredient.

Two consecutive aza-amino acids in peptides promote stable β-turn formation in water.

Studies on the synthetic methodologies and the structural propensity of peptides containing consecutive aza-amino acids are still in their infancy. Here, details of the synthesis and conformational analysis of tripeptides containing two consecutive aza-amino acids are provided. The demonstration that the type I β-turn folding is induced, even in aqueous media, by the introduction of one or two lateral chains on the diaza-peptide unit is of particular importance for the design of peptidomimetics of biological interest.

Polar/apolar interfaces modulate the conformational behavior of cyclic peptides with impact on their passive membrane permeability.

Cyclic peptides have the potential to vastly extend the scope of druggable proteins and lead to new therapeutics for currently untreatable diseases. However, cyclic peptides often suffer from poor bioavailability. To uncover design principles for permeable cyclic peptides, a promising strategy is to analyze the conformational dynamics of the peptides using molecular dynamics (MD) and Markov state models (MSMs). Previous MD studies have focused on the conformational dynamics in pure aqueous or apolar environments to rationalize membrane permeability. However, during the key steps of the permeation through the membrane, cyclic peptides are exposed to interfaces between polar and apolar regions. Recent studies revealed that these interfaces constitute the free energy minima of the permeation process. Thus, a deeper understanding of the behavior of cyclic peptides at polar/apolar interfaces is desired. Here, we investigate the conformational and kinetic behavior of cyclic decapeptides at a water/chloroform interface using unbiased MD simulations and MSMs. The distinct environments at the interface alter the conformational equilibrium as well as the interconversion kinetics of cyclic peptide conformations. For peptides with low population of the permeable conformation in aqueous solution, the polar/apolar interface facilitates the interconversion to the closed conformation, which is required for membrane permeation. Comparison to unbiased MD simulations with a POPC bilayer reveals that not only the conformations but also the orientations are relevant in a membrane system. These findings allow us to propose a permeability model that includes both ‘prefolding’ and ‘non-prefolding’ cyclic peptides – an extension that can lead to new design considerations for permeable cyclic peptides.

Importance of Solvent in Guiding the Conformational Properties of an Intrinsically Disordered Peptide.

Aggregated form of α-synuclein in the brain has been found to be the major component of Lewy bodies that are hallmarks of Parkinson's disease (PD), the second most devastating neurodegenerative disorder. We have carried out room-temperature all-atom molecular dynamics (MD) simulations of an ensemble of widely different α-synuclein1-95 peptide monomer conformations in aqueous solution. Attempts have been made to obtain a generic understanding of the local conformational motions of different repeat unit segments, namely R1-R7, of the peptide and the correlated properties of the solvent at the interface. The analyses revealed relatively greater rigidity of the hydrophobic R6 unit as compared to the other repeat units of the peptide. Besides, water molecules around R6 have been found to be less structured and weakly interacting with the peptide. These are important observations as the R6 unit with reduced conformational motions can act as the nucleation site for the aggregation process, while less structured weakly interacting water around it can become displaced easily, thereby facilitating the hydrophobic collapse of the peptide monomers and their association during the nucleation phase at higher concentrations. In addition, we demonstrated presence of doubly coordinated highly ordered as well as triply coordinated relatively disordered water molecules at the interface. We believe that while the ordered water molecules can favor water-mediated interactions between different peptide monomers, the randomly ordered ones on the other hand are likely to be expelled easily from the interface, thereby facilitating direct peptide-peptide interactions during the aggregation process.

Colanic Acid: Biosynthetic Overproduction by Engineering Escherichia coli and Physical Property Characterization.

Colanic acid has promising applications in food, cosmetic, and healthcare fields. In this study, a recombinant WQM003/pRAU was constructed by deleting genes lon and hns and overexpressing genes rcsA and galU in E. coli MG1655Δ(L-Q). After systematic optimization of fermentation conditions, colanic acid yield in WQM003/pRAU reached 19.79 g/L, the highest yield reported so far. The colanic acid produced by WQM003/pRAU was purified and its structure and physical properties were determined. This colanic acid shows a triple-helical structure and is stable up to 102 °C, and its melting temperature is 253.9 °C. This colanic acid shows a sphere-like chain conformation in aqueous solution. The viscosity of this colanic acid solution is related to concentration, shear rate, salt, temperature, and pH. At high concentrations, this colanic acid shows both viscous and elastic behaviors. These results suggest that the colanic acid produced by WQM003/pRAU has broad application prospects.

Acridine–O6-benzylguanine hybrids: Synthesis, DNA binding, MGMT inhibition and antiproliferative activity

O6-Methylguanine-DNA-methyltransferase (MGMT) is a key DNA repair enzyme involved in chemoresistance to DNA-alkylating anti-cancer drugs such as Temozolomide (TMZ) through direct repair of drug-induced O6-methylguanine residues in DNA. MGMT substrate analogues, such as O6-benzylguanine (BG), efficiently inactivate MGMT in vitro and in cells; however, these drugs failed to reach the clinic due to adverse side effects. Here, we designed hybrid drugs combining a BG residue covalently linked to a DNA-interacting moiety (6-chloro-2-methoxy-9-aminoacridine). Specifically, two series of hybrids, encompassing three compounds each, were obtained by varying the position of the attachment point of BG (N9 of guanine vs. the benzyl group) and the length and nature of the linker. UV/vis absorption and fluorescence data indicate that all six hybrids adopt an intramolecularly stacked conformation in aqueous solutions in a wide range of temperatures. All hybrids interact with double-stranded DNA, as clearly evidenced by spectrophotometric titrations, without intercalation of the acridine ring and do not induce thermal stabilization of the duplex. All hybrids, as well as the reference DNA intercalator (6-chloro-2-methoxy-9-aminoacridine 8), irreversibly inhibit MGMT in vitro with variable efficiency, comparable to that of BG. In a multidrug-resistant glioblastoma cell line T98G, benzyl-linked hybrids 7a–c and the N9-linked hybrid 19b are moderately cytotoxic (GI50 ≥ 15 μM after 96 h), while N9-linked hybrids 19a and 19c are strongly cytotoxic (GI50 = 1–2 μM), similarly to acridine 8 (GI50 = 0.6 μM). Among all compounds, hybrids 19a and 19c, similarly to BG, display synergic cytotoxic effect upon co-treatment with subtoxic doses of TMZ, with combination index (CI) values as low as 0.2–0.3. In agreement with in vitro results, compound 19a inactivates cellular MGMT but, unlike BG, does not induce significant levels of DNA damage, either alone or in combination with TMZ, as indicated by the results of γH2AX immunostaining experiments. Instead, and unlike BG, compound 19a alone induces significant apoptosis of T98G cells, which is not further increased in a combination with TMZ. These results indicate that molecular mechanisms underlying the cytotoxicity of 19a and its combination with TMZ are distinct from that of BG. The strongly synergic properties of this combination represent an interesting therapeutic opportunity in treating TMZ-resistant cancers.

Excited states of six oxazine 1 conformers in aqueous solution: TD-DFT/DFT study

The paper analyzes the electronic-vibrational states of six possible conformers of the dye oxazine 1 (also known as oxazine 725) in an aqueous solution using the SMD solvent model and the 6-31++G(d,p) basis set. Vibronic absorption spectra are calculated using a wide range of hybrid functionals. The X3LYP functional gives the best agreement with the experiment for the positions of the main maximum and shoulder. According to the calculations, the shoulder in the experimental spectrum is of vibronic origin. Various characteristics of the ground and excited states (Duschinsky matrices, IR spectra, atomic charges, dipole moments, maps of the distribution of the electrostatic potential) were obtained. All six investigated conformers have similar vibronic absorption spectra and frontier molecular orbitals. Significant differences between them are observed only for the set of vibronic transitions and relaxation from nonequilibrium to an equilibrium excited state. The effect of two strongly bound water molecules on the excitation of the dye was analyzed.