Specific Side Chains Research Articles

Leaching-Free durable antibacterial composite films constructed via delicate balance between potent contact sterilization and low adhesion

Durable antibacterial surfaces are often required high bactericidal ability and low bacterial adhesion property, but the potent bactericidal surfaces, especially based on the contact bactericidal mechanism, are often paradoxically intended to attract bacteria to adhere. In this work, a delicate balance between the potent contact bactericidal action and low bacterial adhesion was successfully achieved in quaternized fluoropolymer/SiO2 composite films (QFP/SiO2 CFs) through judicious selection of a common fluorinated monomer, a common quaternary ammonium monomer, and a common inert inorganic nanoparticle, to acquire suitable surface energy and chain mobility to ensure low bacterial adhesion, as well as to allow appropriate surface-exposed quaternary ammonium units to synergistically enhance the contact bactericidal action with the specific fluorinated side chains. The QFP/SiO2 CFs were conveniently fabricated through miniemulsion polymerization and followed squeegeeing technique. The QFP/SiO2 CF with an optimized composition exhibited extremely low initial adhesion of bacteria, potent contact bactericidal action, superior long-term antibacterial performance, easy sonication-assistant regeneration for multiple usage, and low cytotoxicity with non-leachable attribute. This work may guide the design and efficient fabrication of leaching-free durable antibacterial surfaces for biological applications, as well as provide an effective strategy to balance contact bactericidal action and low bacteria adhesion for durable antibacterial performance.

Cell penetration of oxadiazole-containing macrocycles.

Passive membrane permeability is an important property in drug discovery and biological probe design. To elucidate the cell-penetrating ability of oxadiazole-containing (Odz) peptides, we employed the Chloroalkane Penetration Assay. The present study demonstrates that Odz cyclic peptides can be highly cell-penetrant depending on the position of specific side chains and the chloroalkane tag. Solution NMR shows that Odz cyclic peptides adopt a β-turn conformation. However, despite observing high cell penetration, we observed low passive permeability in experiments with artificial membranes. These findings highlight the complexity of controlling cell penetration for conformationally sensitive macrocycles and suggest that Odz cyclic peptides may provide a framework for designing cell-penetrant cyclic peptides.

The modification of nisin with homocysteine thiolactone and its effect on antimicrobial activity.

The aim of the present study is to make an important contribution to the literature by focusing on the preparation of the N-homocysteine conjugate of nisin and evaluating the effect of the N-homocysteinylation reaction on its antimicriobial activity. The modification process was monitored using both acetic acid urea polyacrylamide gel electrophoresis (AAU-PAGE) and tricine sodium dodecyl sulphate polyacrylamide gel electrophoresis (tricine SDS-PAGE). The antibacterial effectiveness of modified nisin was assessed against Staphylococcus aureus ATCC 6538, Enterococcus faecium ATCC 9097, Bacillus subtilis ATCC 6633, Lactococcus lactis ssp. cremoris AÜ, Listeria monocytogenes NCTC 5348, and Escherichia coli RSKK. Optimal conditions for achieving the highest N-homocysteinylation degree (6.30%) were determined as 6 mg/mL nisin, 150 mM homocysteine thiolactone, 150 rpm shaking rate, pH of 3.0, and a reaction time of 6 h. The modified nisin obtained did not have a significant inhibitory effect on the strains tested except E. faecium. E. faecium was inhibited by the modified nisin and its antibacterial activity was determined as approximately 10% of the antibacterial activity of unmodified nisin. On the other hand, hydrolysis of nisin by trypsin and thermolysin resulted in significant specific side chain modifications induced by the homocysteine-thiolactone reaction, especially at Lys12 and Lys22. The results provide valuable insights into the potential of N-homocysteinylation to improve the antibacterial properties of nisin and also suggest that the effects of specific modifications identified during the modification process should be investigated.

Xeno Amino Acids: A Look into Biochemistry as We Do Not Know It.

Would another origin of life resemble Earth's biochemical use of amino acids? Here, we review current knowledge at three levels: (1) Could other classes of chemical structure serve as building blocks for biopolymer structure and catalysis? Amino acids now seem both readily available to, and a plausible chemical attractor for, life as we do not know it. Amino acids thus remain important and tractable targets for astrobiological research. (2) If amino acids are used, would we expect the same L-alpha-structural subclass used by life? Despite numerous ideas, it is not clear why life favors L-enantiomers. It seems clearer, however, why life on Earth uses the shortest possible (alpha-) amino acid backbone, and why each carries only one side chain. However, assertions that other backbones are physicochemically impossible have relaxed into arguments that they are disadvantageous. (3) Would we expect a similar set of side chains to those within the genetic code? Many plausible alternatives exist. Furthermore, evidence exists for both evolutionary advantage and physicochemical constraint as explanatory factors for those encoded by life. Overall, as focus shifts from amino acids as a chemical class to specific side chains used by post-LUCA biology, the probable role of physicochemical constraint diminishes relative to that of biological evolution. Exciting opportunities now present themselves for laboratory work and computing to explore how changing the amino acid alphabet alters the universe of protein folds. Near-term milestones include: (a) expanding evidence about amino acids as attractors within chemical evolution; (b) extending characterization of other backbones relative to biological proteins; and (c) merging computing and laboratory explorations of structures and functions unlocked by xeno peptides.

Hyaluronan-arginine enhanced and dynamic interaction emerges from distinctive molecular signature due to electrostatics and side-chain specificity

Hyaluronan is a natural carbohydrate polymer with a negative charge that fosters gel-like conditions crucial for its cellular functions and industrial applications. As a recognized ligand for proteins, understanding their mutual interactions provides solid ground to tune hyaluronan's gel properties using biocompatible peptides. This work employs NMR and molecular dynamics simulations to identify molecular motifs relevant to hyaluronan–peptide interactions using arginine, lysine, and glycine oligopeptides. Arginine-rich peptides exhibit the strongest binding to hyaluronan according to chemical shift perturbation measurements, followed distantly by the similarly charged lysine. This difference highlights the significance of electrostatics and the peculiarities of the guanidinium side chain in arginine, capable of non-polar interactions that further stabilize the binding. Additional nuclear Overhauser effect measurements do not show stable interaction partners, precluding strong and well-defined complexes. Finally, molecular simulations support our findings and show an extended but significant interaction region, especially for arginine, responsible for the observed enhanced binding, which can also promote cross-linking of hyaluronan polymers. Our findings pave the way for optimizing biocompatible peptides to alter hyaluronan gels' properties efficiently and also explain why hyaluronan–protein interaction typically involves positively charged arginine-rich regions also capable of forming hydrogen bonds and non-polar interactions.

Controlling the Chameleonic Behavior and Membrane Permeability of Cyclosporine Derivatives via Backbone and Side Chain Modifications.

Some macrocycles exhibit enhanced membrane permeability through conformational switching in different environmental polarities, a trait known as chameleonic behavior. In this study, we demonstrate specific backbone and side chain modifications that can control chameleonic behavior and passive membrane permeability using a cyclosporin O (CsO) scaffold. To quantify chameleonic behavior, we used a ratio of the population of the closed conformation obtained in polar solvent and nonpolar solvent for each CsO derivative. We found that β-hydroxylation at position 1 (1 and 3) can encode chameleonicity and improve permeability. However, the conformational stabilization induced by adding an additional transannular H-bond (2 and 5) leads to a much slower rate of membrane permeation. Our CsO scaffold provides a platform for the systematic study of the relationship among conformation, membrane permeability, solubility, and protein binding. This knowledge contributes to the discovery of potent beyond the rule of five (bRo5) macrocycles capable of targeting undruggable targets.

PARP14 is a writer, reader, and eraser of mono-ADP-ribosylation

PARP14/BAL2 is a large multidomain enzyme involved in signaling pathways with relevance to cancer, inflammation, and infection. Inhibition of its mono-ADP-ribosylating PARP homology domain and its three ADP-ribosyl binding macro domains has been regarded as a potential means of therapeutic intervention. Macrodomains-2 and -3 are known to stably bind to ADP-ribosylated target proteins, but the function of macrodomain-1 has remained somewhat elusive. Here, we used biochemical assays of ADP-ribosylation levels to characterize PARP14 macrodomain-1 and the homologous macrodomain-1 of PARP9. Our results show that both macrodomains display an ADP-ribosyl glycohydrolase activity that is not directed toward specific protein side chains. PARP14 macrodomain-1 is unable to degrade poly(ADP-ribose), the enzymatic product of PARP1. The F926A mutation of PARP14 and the F244A mutation of PARP9 strongly reduced ADP-ribosyl glycohydrolase activity of the respective macrodomains, suggesting mechanistic homology to the Mac1 domain of the SARS-CoV-2 Nsp3 protein. This study adds two new enzymes to the previously known six human ADP-ribosyl glycohydrolases. Our results have key implications for how PARP14 and PARP9 will be studied and how their functions will be understood.

Hydroxy Acid Activation in Fungal Non-Ribosomal Peptide Synthesis Assessed by Site-Directed Mutagenesis.

The fungal cyclodepsipeptides (CDPs) enniatin, beauvericin, bassianolide, and PF1022 consist of alternating N-methylated l-amino and d-hydroxy acids. They are synthesized by non-ribosomal peptide synthetases (NRPS). The amino acid and hydroxy acid substrates are activated by adenylation (A) domains. Although various A domains have been characterized thus giving insights into the mechanism of substrate conversion, little is known about the utilization of hydroxy acids in NRPSs. Therefore, we used homology modelling and molecular docking of the A1 domain of enniatin synthetase (EnSyn) to gain insights into the mechanism of hydroxy acid activation. We introduced point mutations into the active site and used a photometric assay to study the substrate activation. The results suggest that the hydroxy acid is selected by interaction with backbone carbonyls rather than by a specific side chain. These insights enhance the understanding of non-amino acid substrate activation and could contribute to the engineering of depsipeptide synthetases.

Emergence of Cooperative Glucose-Binding Networks in Adaptive Peptide Systems.

Minimalistic peptide-based systems that bind sugars in water are challenging to design due to the weakness of interactions and required cooperative contributions from specific amino-acid side chains. Here, we used a bottom-up approach to create peptide-based adaptive glucose-binding networks by mixing glucose with selected sets of input dipeptides (up to 4) in the presence of an amidase to enable in situ reversible peptide elongation, forming mixtures of up to 16 dynamically interacting tetrapeptides. The choice of input dipeptides was based on amino-acid abundance in glucose-binding sites found in the protein data bank, with side chains that can support hydrogen bonding and CH-π interactions. Tetrapeptide sequence amplification patterns, determined through LC-MS analysis, served as a readout for collective interactions and led to the identification of optimized binding networks. Systematic variation of dipeptide input revealed the emergence of two networks of non-covalent hydrogen bonding and CH-π interactions that can co-exist, are cooperative and context-dependent. A cooperative binding mode was determined by studying the binding of the most amplified tetrapeptide (AWAD) with glucose in isolation. Overall, these results demonstrate that the bottom-up design of complex systems can recreate emergent behaviors driven by covalent and non-covalent self-organization that are not observed in reductionist designs and lead to the identification of system-level cooperative binding motifs.

Towards high-throughput screening (HTS) of polyhydroxyalkanoate (PHA) production via Fourier transform infrared (FTIR) spectroscopy of Halomonas sp. R5-57 and Pseudomonas sp. MR4-99.

High-throughput screening (HTS) methods for characterization of microbial production of polyhydroxyalkanoates (PHA) are currently under investigated, despite the advent of such systems in related fields. In this study, phenotypic microarray by Biolog PM1 screening of Halomonas sp. R5-57 and Pseudomonas sp. MR4-99 identified 49 and 54 carbon substrates to be metabolized by these bacteria, respectively. Growth on 15 (Halomonas sp. R5-57) and 14 (Pseudomonas sp. MR4-99) carbon substrates was subsequently characterized in 96-well plates using medium with low nitrogen concentration. Bacterial cells were then harvested and analyzed for putative PHA production using two different Fourier transform infrared spectroscopy (FTIR) systems. The FTIR spectra obtained from both strains contained carbonyl-ester peaks indicative of PHA production. Strain specific differences in the carbonyl-ester peak wavenumber indicated that the PHA side chain configuration differed between the two strains. Confirmation of short chain length PHA (scl-PHA) accumulation in Halomonas sp. R5-57 and medium chain length PHA (mcl-PHA) in Pseudomonas sp. MR4-99 was done using Gas Chromatography-Flame Ionization Detector (GC-FID) analysis after upscaling to 50 mL cultures supplemented with glycerol and gluconate. The strain specific PHA side chain configurations were also found in FTIR spectra of the 50 mL cultures. This supports the hypothesis that PHA was also produced in the cells cultivated in 96-well plates, and that the HTS approach is suitable for analysis of PHA production in bacteria. However, the carbonyl-ester peaks detected by FTIR are only indicative of PHA production in the small-scale cultures, and appropriate calibration and prediction models based on combining FTIR and GC-FID data needs to be developed and optimized by performing more extensive screenings and multivariate analyses.

Molecular fingerprint in hyaluronan-peptide interactions: Cooperation of electrostatics and side-chain specificity in enhanced arginine effect.

Hyaluronan is a sugar polymer found on the extracellular side, close to the plasma membrane. It is the scaffold of the extracellular matrix due to its high solubility, despite its low negative charge density and unusual length. Hyaluronan is also a known ligand of several glycan-binding proteins. We can improve our understanding of these interactions by characterizing hyaluronan affinity to model amino acid sequences of reduced complexity. Extracted features and fingerprints allow for identifying new potential binding partners and expand the current understanding of hyaluronan's biological role. Here, we use a combination of NMR and molecular dynamics simulations (MD) to study hyaluronan binding to short oligopeptides. We focus on arginine as a known critical amino acid for this interaction and compare it to lysine and glycine to highlight its distinctive features. We can test the binding using 1H NMR chemical shift perturbation (CSP) by titrating hyaluronan with a peptide. CSP not only estimates binding strength but also pinpoints the binding motif involved. Our findings are further explained by MD, which provide an atomistic fingerprint of the hyaluronan-peptide complex. Interestingly, we could not find any NOE signal confirming the CSP observed interactions. The combination of positive CSP with no visible NOEs indicates the presence of a dynamic binding mode in agreement with our MD simulations. This agreement with simulations was only achieved using the prosECCo force field for accounting for electronic polarizability. Overall, we find that arginine, a positively charged amino acid, binds the strongest to the negative hyaluronan, followed by the positive lysine. The hyaluronan's carboxyl and amide groups are the driving force for interacting with the peptides. We prove the importance of the electrostatics interaction but also a specific sidechain affinity that favors arginine.

A Transfer Free Energy Based Implicit Solvent Model for Protein Simulations in Solvent Mixtures: Urea-Induced Denaturation as a Case Study.

We developed a method for implicit solvent molecular dynamics simulations of proteins in solvent mixtures (model with implicit solvation thermodynamics, MIST). The MIST method introduces experimental group transfer free energies to the generalized Born formulation for generating molecular trajectories without the need for developing rigorous explicit-solvent force fields for multicomponent solutions. As a test case, we studied the urea-induced denaturation of the Trp-cage miniprotein in water. We demonstrate that our method allows efficient exploration of the conformational space of the protein in only a few hundreds of nanoseconds of all-atom unbiased simulations. Furthermore, selective implementation of the transfer free energies of specific peptide groups, backbone, and side chains enables us to decouple their specific energetic contributions to the conformational changes of the protein. The approach herein developed can readily be extended to the investigation of complex matrices as well as to the characterization of protein aggregation. The MIST method is implemented in Plumed (ver. 2.8) as a separate module called SASA.

A Computational Analysis of the Reaction of SO2 with Amino Acid Anions: Implications for Its Chemisorption in Biobased Ionic Liquids

We report a series of calculations to elucidate one possible mechanism of SO2 chemisorption in amino acid-based ionic liquids. Such systems have been successfully exploited as CO2 absorbents and, since SO2 is also a by-product of fossil fuels’ combustion, their ability in capturing SO2 has been assessed by recent experiments. This work is exclusively focused on evaluating the efficiency of the chemical trapping of SO2 by analyzing its reaction with the amino group of the amino acid. We have found that, overall, SO2 is less reactive than CO2, and that the specific amino acid side chain (either acid or basic) does not play a relevant role. We noticed that bimolecular absorption processes are quite unlikely to take place, a notable difference with CO2. The barriers along the reaction paths are found to be non-negligible, around 7–11 kcal/mol, and the thermodynamic of the reaction appears, from our models, unfavorable.

Strategies to Control the Cis-Trans Isomerization of Peptoid Amide Bonds.

Peptoids are oligomers of N-substituted glycine units. They structurally resemble peptides but, unlike natural peptides, the side chains of peptoids are present on the amide nitrogen atoms instead of the α-carbons. The N-substitution improves cell-permeability of peptoids and enhance their proteolytic stability over natural peptides. Therefore, peptoids are ideal peptidomimetic candidates for drug discovery, especially for intracellular targets. Unfortunately, most peptoid ligands discovered so far possess moderate affinity towards their biological targets. The moderate affinity of peptoids for biomacromolecules is linked to their conformational flexibility, which causes substantial entropic loss during the peptoid-biomacromolecule binding process. The conformational flexibility of peptoids is caused by the lack of backbone chirality, absence of hydrogen bond donors (NH) in their backbone to form CO⋅⋅⋅HN hydrogen bonds and the facile cis-trans isomerization of their tertiary amide bonds. In recent years, many investigators have shown that the incorporation of specific side chains with unique steric and stereoelectronic features can favourably shift the cis-trans equilibria of peptoids towards one of the two isomeric forms. Such strategies are helpful to design homogenous peptoid oligomers having well defined secondary structures. Herein, we discuss the strategies developed over the years to control the cis-trans isomerization of peptoid amide bonds.

Pegylation of the Galectin-3 carbohydrate recognition domain creates a kinetic trap protecting the protein from thermal unfolding

Proteins are widely used as therapeutics but often can be unstable, limiting their applications in the clinic. To improve the stability of protein drugs, synthetic polymers such as polyethylene glycol (PEG) can be covalently attached to specific protein side chains, creating protein-polymer conjugates. Though nearly thirty PEGylated proteins have been FDA-approved, our understanding is very limited of how PEG-protein interactions alter the fundamental chemical and physical properties of the conjugate.

Gas-phase formation of cationic fullerene/9-aminoanthracene clusters: an indicator for interstellar dust growth

ABSTRACT Growth of clusters by adduction of monomers – as the first step in dust particle growth – is an area of much interest in astronomy. We focus on the fullerene/9-aminoanthracene cluster species, to illustrate the competition between the van der Waals bonding growth and the covalent bonding growth model versus the charge transfer model in the large cluster formation process. The experimental results show that fullerene-fragment (C56 and C58)/9-aminoanthracene cluster cations, e.g. [(C14H11N)nC56]+ and [(C14H11N)nC58]+, n = [1,7], are efficiently formed, while C$_{60}^+$ is insensitive to the cluster’s formation. With laser irradiation, all the fullerene/9-aminoanthracene clusters dissociate into 9-aminoanthracene and fullerene cations. The mechanisms for the reactions of fullerene cations and 9-aminoanthracene were investigated by theoretical calculations, under the assumption that the molecular geometries found for the formed complexes correspond to the global energy minima: the absence of C$_{60}^+$ clusters is mainly due to the charge transfer channel’s competition; [(C14H11N)C58]+ has three types of isomers, with van der Waals or covalent bonds, mainly depending on the reaction sites of fullerene cations. Importantly, in the size grown process, for the fullerene/9-aminoanthracene cluster there exists a geometry configuration conversion between the van der Waals and covalent bonding modes. The largest fullerene/9-aminoanthracene clusters, e.g. [(C14H11N)7C58]+ (240 atoms, ∼4 nm in size), are likely in a multishelled geometry, i.e. seven 9-aminoanthracene molecules surrounding fullerene cations in two layers, which can directly build the relationship between molecular clusters and carbonaceous grains. Nitrogen matters! The specific side chains (e.g. –NH2) play an important role in the growth of interstellar dust.

CIMAGE2.0: An Expanded Tool for Quantitative Analysis of Activity-Based Protein Profiling (ABPP) Data.

Activity-based protein profiling (ABPP) is a powerful chemical proteomic method for studying protein activity, modifications, and interactions in a high-throughput manner. In ABPP experiments, accurate quantification is crucial to determine the extent of probe labeling at the level of either target proteins or specific amino acid side chains. CIMAGE has been developed as an in-house quantification software specifically designed for ABPP data analysis that incorporates (1) a relaxed peak extraction algorithm and (2) stringent post-quantification checks for efficient and accurate quantification. It also can generate table and image data for users to conveniently visualize their results. Here we provide a retrospective introduction of the software and describe our recent upgrade efforts to enable (1) interfacing with different database search engines as input, (2) triplex quantification of ABPP data by reductive dimethylation, and (3) envelope checking for chemical elements with special isotopic distributions. We show that the updated CIMAGE can maintain its ability to quantify ABPP data with dramatic depth and high accuracy, and it also has similar quantification performance in benchmarked SILAC data as compared with MaxQuant. We believe that CIMAGE2.0 will continue to serve as a powerful analytical tool for ABPP studies.

Recent Advances in Molecular Design of Organic Thermoelectric Materials

Recent Advances in Molecular Design of Organic Thermoelectric Materials

Molecular Brush Surfactants: Versatile Emulsifiers for Stabilizing and Structuring Liquids

AbstractUsing amphiphilic molecular brushes to stabilize emulsions usually requires the synthesis of specific side chains, which can be a time‐consuming and difficult challenge to meet. By taking advantage of the electrostatic interactions between water‐soluble molecular brushes and oil‐soluble oligomeric ligands, the in situ formation, assembly and jamming of molecular brush surfactants (MBSs) at the oil‐water interface is described. With MBSs, stable emulsions including o/w, w/o and o/w/o can be easily prepared by varying the molar ratios of the molecular brushes to the ligands. Moreover, when jammed, the binding energy of MBSs at the interface is sufficiently strong to allow the stabilization of liquids in nonequilibrium shapes, i.e., structuring liquids, producing an elastic film at the interface with exceptional mechanical properties. These structured liquids have numerous potential applications, including chemical biphasic reactions, liquid electronics, and all‐liquid biomimetic system.

Molecular Brush Surfactants: Versatile Emulsifiers for Stabilizing and Structuring Liquids.

Using amphiphilic molecular brushes to stabilize emulsions usually requires the synthesis of specific side chains, which can be a time-consuming and difficult challenge to meet. By taking advantage of the electrostatic interactions between water-soluble molecular brushes and oil-soluble oligomeric ligands, the in situ formation, assembly and jamming of molecular brush surfactants (MBSs) at the oil-water interface is described. With MBSs, stable emulsions including o/w, w/o and o/w/o can be easily prepared by varying the molar ratios of the molecular brushes to the ligands. Moreover, when jammed, the binding energy of MBSs at the interface is sufficiently strong to allow the stabilization of liquids in nonequilibrium shapes, i.e., structuring liquids, producing an elastic film at the interface with exceptional mechanical properties. These structured liquids have numerous potential applications, including chemical biphasic reactions, liquid electronics, and all-liquid biomimetic system.