Side Chain Hydroxyl Research Articles

Indirect Formation of Peptide Bonds as a Prelude to Ribosomal Transpeptidation.

The catalytic competency of the ribosome in extant protein biosynthesis is thought to arise primarily from two sources: an ability to precisely juxtapose the termini of two key substrates─3'-aminoacyl and N-acyl-aminoacyl tRNAs─and an ability to ease direct transpeptidation by their desolvation and encapsulation. In the absence of ribosomal, or enzymatic, protection, however, these activated alkyl esters undergo efficient hydrolysis, while significant entropic barriers serve to hamper their intermolecular cross-aminolysis in bulk water. Given that the spontaneous emergence of a catalyst of comparable size and sophistication to the ribosome in a prebiotic RNA world would appear implausible, it is thus natural to ask how appreciable peptide formation could have occurred with such substrates in bulk water without the aid of advanced ribozymatic catalysis. Using a combination of fluorine-tagged aminoacyl adenylate esters, in situ monitoring by 19F{1H} NMR spectroscopy, analytical deconvolution of kinetics, pH-rate profile analysis, and temperature-dependence studies, we here explore the mechanistic landscape of indirect amidation, via transesterification and O-to-N rearrangement, as a highly efficient, alternative manifold for transpeptidation that may have served as a prelude to ribosomal peptide synthesis. Our results suggest a potentially overlooked role for those amino acids implicated by the cyanosulfidic reaction network with hydroxyl side chains (Ser and Thr), and they also help to resolve some outstanding ambiguities in the broader literature regarding studies of similar systems (e.g., aminolyzes with Tris buffer). The evolutionary implications of this mode of peptide synthesis and the involvement of a very specific subset of amino acids are discussed.

Self-contacting Cys, Ser, and Thr residues in high-resolution protein crystal structures: Tertiary constraints or hydrogen bonds?

Functional groups in the side-chains of at least 10 amino acids are mainly involved in tertiary interactions. However, structural and functional significance of intra-residue interactions has not been fully recognized. In this study, we have analyzed ~5800 non-redundant high-resolution protein structures and identified 1166 self-contacts between the side-chain S-H/O-H and backbone C=O groups in Cys, Ser, and Thr residues that satisfied the geometric criteria to form hydrogen bonds. Quantum chemical calculations using model compounds were used to evaluate single point energy for 45 representative examples from different allowed regions of Ramachandran map. Relative energy profiles obtained by varying the side-chain dihedral angle χ1 revealed that the energy difference between the crystal structure and the minimum energy conformations is between 0 and 3 kcal/mol. Natural bond orbital analysis (NBO) of self-contacting Cys residues revealed no charge transfer between Cys side-chain S-H and the backbone C=O groups. However, side-chain hydroxyl and the backbone C=O groups of 90%-95% of all self-contacting Ser and Thr residues are involved in charge transfer and the second order perturbation energy of majority of them is above 1 kcal/mol. Interaction energies calculated for model compounds along with NBO and NCIPLOT analyses demonstrate that the self-contacts observed in Ser and Thr residues can be described as hydrogen bonds. These interactions may provide stability to the loop/coil conformations. Self-contacting Cys residues are buried and the self-contacts appear to be mostly due to tertiary constraints. Dispersion between the self-contacting groups is one way to explain the close approach in Cys residues. Mutation studies will further validate and reveal the structural and functional significance of these self-contacting residues.

Ease-of-manufacture highly transparent thin polyvinyl alcohol aerogel

The earliest silicon-based aerogels attracted attention due to their nanoscale porous structure and high transparency. Still, they need to be more balanced with the poor mechanical properties. Their brittle structure limits the development of this promising new material. Therefore, the goal of this work is to optimize the mechanical properties of aerogels while maintaining transparency. The good mechanical properties of polymers have made them the material of choice for this work. Polyvinyl alcohol (PVA), which can undergo self-crosslinking through side-chain hydroxyl groups forming hydrogen bonds, was chosen as the raw material to simplify and expedite the production process. The production process was experimented with, analyzed, and refined. Considering the time efficiency of the experimental process, a one-step gelling method was invented to facilitate the sol-gel transition. The one-step gelling method maintained the high transparency of PVA aerogels and reduced the time required for the gelation process compared to the freeze-thawing method. This work employs carbon dioxide supercritical drying to ensure minimal structural collapse and maximize the porous structure’s retention. The transmittance of PVA aerogels can reach up to 93.67% at a wavelength of 1333 nm. The internal structure of aerogels with different PVA concentrations was observed using a Scanning electron microscope. Applying Beer-Lambert’s Law eliminated the effect of sample thickness on transparency. The relationship between the transparency of PVA aerogels, their microstructure, and macroscopic concentration was studied and analyzed for the first time. While ensuring light transmittance, the modulus of the PVA aerogel reached as high as 6.18 ± 0.56 MPa at 13 wt%.

In Situ Functionalization of Polar Polythiophene-Based Organic Electrochemical Transistor to Interface In Vitro Models.

Organic mixed ionic-electronic conductors are promising materials for interfacing and monitoring biological systems, with the aim of overcoming current challenges based on the mismatch between biological materials and convectional inorganic conductors. The conjugated polymer/polyelectrolyte complex poly(3,4-ethylenedioxythiophene):polystyrenesulfonate (PEDOT/PSS) is, up to date, the most widely used polymer for in vitro or in vivo measurements in the field of organic bioelectronics. However, PEDOT/PSS organic electrochemical transistors (OECTs) are limited by depletion mode operation and lack chemical groups that enable synthetic modifications for biointerfacing. Recently introduced thiophene-based polymers with oligoether side chains can operate in accumulation mode, and their chemical structure can be tuned during synthesis, for example, by the introduction of hydroxylated side chains. Here, we introduce a new thiophene-based conjugated polymer, p(g42T-T)-8% OH, where 8% of the glycol side chains are functionalized with a hydroxyl group. We report for the first time the compatibility of conjugated polymers containing ethylene glycol side chains in direct contact with cells. The additional hydroxyl group allows covalent modification of the surface of polymer films, enabling fine-tuning of the surface interaction properties of p(g42T-T)-8% OH with biological materials, either hindering or promoting cell adhesion. We further use p(g42T-T)-8% OH to fabricate the OECTs and demonstrate for the first time the monitoring of epithelial barrier formation of Caco-2 cells in vitro using accumulation mode OECTs. The conjugated polymer p(g42T-T)-8% OH allows organic-electronic-based materials to be easily modified and optimized to interface and monitor biological systems.

Exploring the reactivity and interactions of a poly(fluorene‐co‐tetrazine)‐conjugated polymer with Single‐walled Carbon Nanotubes

AbstractConjugated tetrazine‐containing polymers that undergo inverse electron demand Diels‐Alder (IEDDA) reactions with trans‐cyclooctenes are interesting not only for their intrinsic optoelectronic properties, but also their interactions with π‐conjugated surfaces. Here, we prepared a series of poly(fluorene‐co‐tetrazine) polymers and carried out IEDDA reactions to decorate them with hydroxyl, hexadecyl, or triethylene glycol side chains. The polymers were investigated pre‐ and post‐IEDDA coupling in terms of their ability to disperse single‐walled carbon nanotubes (SWNTs) in organic solvent. It was found that polymer molecular weight, side chain structure, and degree of conjugation all impacted the quality of SWNT dispersions. While the starting poly(fluorene‐co‐tetrazine) polymer produced concentrated dispersions, the post‐IEDDA polymer containing dihydropyridazine groups did not produce dispersions of equal concentration. However, upon oxidation to the fully aromatic pyridazines, the polymers regained their ability to form concentrated dispersions. Furthermore, the post‐IEDDA polymers exhibited increased selectivity toward metallic SWNTs relative to the starting polymer. Due to the efficiency of the IEDDA reaction, it was also possible to use this chemistry to derivatize the nanotube complex with poly(fluorene‐co‐tetrazine) post‐dispersion. Overall, this work demonstrates the first use of reactive polytetrazines to disperse SWNTs, allowing rapid modification of polymer‐nanotube complexes.

Utilizing Thermal Shift Assay to Probe Substrate Binding to Selenoprotein O.

Defining the biological importance of proteins with unknown functions poses a significant obstacle in understanding cellular processes. Although bioinformatic and structural predictions have contributed to the study of unknown proteins, in vitro experimental validations are often hampered by the optimal conditions and cofactors required for biochemical activity. Cofactor binding is not only essential for the activity of some enzymes but may also enhance the thermal stability of the protein. One practical application of this phenomenon lies in utilizing the change in thermal stability, as measured by alterations in the protein's melting temperature, to probe ligand binding. Thermal shift assay (TSA) can be used to analyze the binding of different ligands to the protein of interest or find a stabilizing condition to perform experiments such as X-ray crystallography. Here we will describe a protocol for TSA utilizing the pseudokinase, Selenoprotein O (SelO), for a simple and high-throughput method for testing metal and nucleotide binding. In contrast to canonical kinases, SelO binds ATP in an inverted orientation to catalyze the transfer of AMP to the hydroxyl side chains of proteins in a posttranslational modification known as protein AMPylation. By leveraging the shift in the melting temperatures, we provide crucial insights into the molecular interactions underlying SelO function.

Design and synthesis of novel and potent allosteric HIV-1 integrase inhibitors with a spirocyclic moiety

We report herein the design and discovery of novel allosteric HIV-1 integrase inhibitors. Our design concept utilized the spirocyclic moiety to restrain the flexibility of the conformation of the lipophilic part of the inhibitor. Compound 5 showed antiviral activity by binding to the nuclear lens epithelium-derived growth factor (LEDGF/p75) binding site of HIV-1 integrase (IN). The introduction of a lipophilic amide substituent into the central benzene ring resulted in a significant increase in antiviral activity against HIV-1 WT X-ray crystallography of compound 15 in complex with the integrase revealed the presence of a hydrogen bond between the oxygen atom of the amide of compound 15 and the hydroxyl group of the T125 side chain. Chiral compound 17 showed high antiviral activity, good bioavailability, and low clearance in rats.

Impact of single-residue mutations on protein thermal stability: The case of threonine 83 of BC2L-CN lectin

The thermal stability of trimeric lectin BC2L-CN was investigated and found to be considerably altered when mutating residue 83, originally a threonine, located at the fucose-binding loop. Mutants were analyzed using differential scanning calorimetry and isothermal microcalorimetry. Although most mutations decreased the affinity of the protein for oligosaccharide H type 1, six mutations increased the melting temperature (Tm) by >5 °C; one mutation, T83P, increased the Tm value by 18.2 °C(T83P, Tm = 96.3 °C). In molecular dynamic simulations, the investigated thermostable mutants, T83P, T83A, and T83S, had decreased fluctuations in the loop containing residue 83. In the T83S mutation, the side-chain hydroxyl group of serine formed a hydrogen bond with a nearby residue, suggesting that the restricted movement of the side-chain resulted in fewer fluctuations and enhanced thermal stability. Residue 83 is located at the interface and near the upstream end of the equivalent loop in a different protomer; therefore, fluctuations by this residue likely propagate throughout the loop. Our study of the dramatic change in thermal stability by a single amino acid mutation provides useful insights into the rational design of protein structures, especially the structures of oligomeric proteins.

Conformations of a highly expressed Z19 α-zein studied with AlphaFold2 and MD simulations.

α-zeins are amphiphilic maize seed storage proteins with material properties suitable for a multitude of applications e.g., in renewable plastics, foods, therapeutics and additive manufacturing (3D-printing). To exploit their full potential, molecular-level insights are essential. The difficulties in experimental atomic-resolution characterization of α-zeins have resulted in a diversity of published molecular models. However, deep-learning α-zein models are largely unexplored. Therefore, this work studies an AlphaFold2 (AF2) model of a highly expressed α-zein using molecular dynamics (MD) simulations. The sequence of the α-zein cZ19C2 gave a loosely packed AF2 model with 7 α-helical segments connected by turns/loops. Compact tertiary structure was limited to a C-terminal bundle of three α-helices, each showing notable agreement with a published consensus sequence. Aiming to chart possible α-zein conformations in practically relevant solvents, rather than the native solid-state, the AF2 model was subjected to MD simulations in water/ethanol mixtures with varying ethanol concentrations. Despite giving structurally diverse endpoints, the simulations showed several patterns: In water and low ethanol concentrations, the model rapidly formed compact globular structures, largely preserving the C-terminal bundle. At ≥ 50 mol% ethanol, extended conformations prevailed, consistent with previous SAXS studies. Tertiary structure was partially stabilized in water and low ethanol concentrations, but was disrupted in ≥ 50 mol% ethanol. Aggregated results indicated minor increases in helicity with ethanol concentration. β-sheet content was consistently low (∼1%) across all conditions. Beyond structural dynamics, the rapid formation of branched α-zein aggregates in aqueous environments was highlighted. Furthermore, aqueous simulations revealed favorable interactions between the protein and the crosslinking agent glycidyl methacrylate (GMA). The proximity of GMA epoxide carbons and side chain hydroxyl oxygens simultaneously suggested accessible reactive sites in compact α-zein conformations and pre-reaction geometries for methacrylation. The findings may assist in expanding the applications of these technologically significant proteins, e.g., by guiding chemical modifications.

Molecular engineering of π-extended viologens consisting of oxadiazoles-based bridges for highly stable electrochromic devices

Four π-extended viologen derivatives (Spr-PO/Sbu-PO/OHpr-PO/ OHet-PO) were synthesized by introducing oxadiazole group and side chain modification. Compared with viologen with bipyridines as the chromophore, these POs possessed extended conjugation lengths and optimized photoelectron conversion efficiency. The CMC gel electrochromic devices based on them could realize the color transformation from colorless to yellow-green under the working voltage of 1.1 V. Furthermore, due to different side chain groups, the diffusion rates of POs in the gel changed significantly. Compounds based on hydroxyl side chain groups showed higher response rates (<10 s) in the CMC gel system, while sulfonate side chain modified POs showed slower response characteristics (>10 s). All POs showed excellent stability, the optical contrasts were almost unchanged after continuous working for 100 cycles. This study provided an important method for obtaining efficient and stable electrochromic devices.

Effect of lignin modification on the selectivity of pyrolysis products from softwood kraft lignin

The synthesis of phenol via pyrolysis of lignin can be envisaged as a prospective approach for the high-value utilization of lignin, Herein, we describe an in-depth investigation of the selective modification of lignin to effectively adjust the distribution of lignin pyrolysis products. We found that hydroxypropyl modification of lignin can facilitate the elimination of methoxy from the lignin macromolecule and improve the selectivity of phenol products. In addition, increasing the carbon number of the lignin side chains causes polymerization of pyrolysis products, which is unfavorable for the synthesis of fine chemicals via lignin pyrolysis. Conversely, an increase in lignin side-chain hydroxyl content can lead to more liquid-phase products from lignin pyrolysis, and the selectivity of phenol and its congeners will also increase. As a result, the yield of liquid products obtained via pyrolysis of lignin modified via selective oxyalkylation can reach 52 %, and the relative content of phenol and its homologs can reach 45.2 %.

Protection against β-N-methylamino-l-alanineꟷinduced vesicular monoamine transporter 2 inhibition by hydroxyl-containing proteinogenic amino acids

β-N-methylamino-l-alanine (BMAA) has been shown to inhibit vesicular monoamine transporter 2 (VMAT2), thereby preventing the uptake of monoaminergic neurotransmitters into platelet dense granules and synaptic vesicles. The inhibition is hypothesized to be through direct association of BMAA with hydroxyl groupꟷcontaining amino acid residues in VMAT2. This study evaluated whether BMAA-induced inhibition of VMAT2 could be prevented directly by co-incubation of BMAA with amino acids, and if this protection was specific for BMAA inhibition of VMAT2. l-tyrosine, and to a lesser extent l-serine, was able to prevent BMAA-induced VMAT2 inhibition in a concentration-dependent manner, whereas neither l-threonine nor amino acids without side chain hydroxyl groups could reduce this inhibition. Reserpine-induced VMAT2 inhibition was unaffected by any of the amino acids. These data support the hypothesized interaction between BMAA and hydroxyl groupꟷcontaining amino acids and suggests that this interaction might be leveraged to protect against the toxicity of BMAA.

Construction methods and biomedical applications of PVA-based hydrogels.

Polyvinyl alcohol (PVA) hydrogel is favored by researchers due to its good biocompatibility, high mechanical strength, low friction coefficient, and suitable water content. The widely distributed hydroxyl side chains on the PVA molecule allow the hydrogels to be branched with various functional groups. By improving the synthesis method and changing the hydrogel structure, PVA-based hydrogels can obtain excellent cytocompatibility, flexibility, electrical conductivity, viscoelasticity, and antimicrobial properties, representing a good candidate for articular cartilage restoration, electronic skin, wound dressing, and other fields. This review introduces various preparation methods of PVA-based hydrogels and their wide applications in the biomedical field.

Synthesis and Identification of decarbamoyloxySaxitoxins in Toxic Microalgae and their Reactions with the Oxygenase, SxtT, Reveal Saxitoxin Biosynthesis.

Saxitoxin (STX, 1) is a representative compound of paralytic shellfish toxins (PSTs) that are produced by marine dinoflagellates and freshwater cyanobacteria. Although several pathways have been proposed for the biosynthesis of STX, the order of ring and side chain hydroxylation, and formation of the tricyclic skeleton have not been well established. In this study, 12,12-dideoxy-decarbamoyloxySTX (dd-doSTX, 2), the most reduced STX analogue having the tricyclic skeleton, and its analogues, 12β-deoxy-doSTX (12β-d-doSTX, 3), 12α-deoxy-doSTX (12α-d-doSTX, 4), and doSTX (5), were synthesized, and these compounds were screened in the toxic microalgae using high-resolution LCMSMS. dd-doSTX (2) and 12β-d-doSTX (3) were identified in the PSTs-producing dinoflagellates (Alexandrium catenella, A. pacificum, and/or Gymnodinium catenatum) and in the cyanobacterium Dolichospermum circinale (TA04). doSTX (5), previously isolated from the dinoflagellate G. catenatum, was also identified in D. circinale (TA04). Furthermore, the conversion of 2 to 3, and 4 to 5, by SxtT with VanB, a reported Rieske oxygenase and its redox partner in STX biosynthesis, was confirmed. These results support that 2 is a possible biosynthetic precursor of STX, and that ring and side-chain hydroxylations proceed after cyclization.

Three-Minute Enantioselective Amino Acid Analysis by Ultra-High-Performance Liquid Chromatography Drift Tube Ion Mobility-Mass Spectrometry Using a Chiral Core-Shell Tandem Column Approach.

Fast liquid chromatography (LC) amino acid enantiomer separation of 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatives using a chiral core-shell particle tandem column with weak anion exchange and zwitterionic-type quinine carbamate selectors in less than 3 min was achieved. Enantiomers of all AQC-derivatized proteinogenic amino acids and some isomeric ones (24 in total plus achiral glycine) were baseline separated (Rs > 1.5 except for glutamic acid with Rs = 1.3), while peaks of distinct amino acids and structural isomers (constitutional isomers and diastereomers of leucine and threonine) of the same configuration overlapped to various degrees. For this reason, drift tube ion mobility-mass spectrometry was added (i.e., LC-IM-MS) as an additional selectivity filter without extending run time. The IM separation dimension in combination with high-resolution demultiplexing enabled confirmation of threonine isomers (threonine, allo-threonine, homoserine), while leucine, isoleucine, and allo-isoleucine have almost identical collisional cross-section (DTCCSN2) values and added no selectivity to the partial LC separation. Density functional theory (DFT) calculations show that IM separation of threonine isomers was possible due to conformational stabilization by hydrogen bond formation between the hydroxyl side chain and the urea group. Generally, the CCSN2 of protonated ions increased uniformly with addition of the AQC label, while outliers could be explained by consideration of intramolecular interactions and additional structural analysis. Preliminary validation of the enantioselective LC-IM-MS method for quantitative analysis showed compliance of accuracy and precision with common limits in bioanalytical methods, and applicability to a natural lipopeptide and a therapeutic synthetic peptide could be demonstrated.

Syntheses of optically active monapterin, 7,8-dihydromonapterin, and 5,6,7,8-tetrahydromonapterin from l-xylose.

We describe the syntheses of monapterin, dihydromonapterin and tetrahydromonapterin in optically active forms. The syntheses involved the condensation of l-xylose with phenylhydrazine, providing a hydrazone derivative. The reaction of the resulting hydrazone with triamino-pyrimidinone followed by oxidation of the resulting pteridinone with molecular oxygen furnished pterin containing a hydroxylated side chain. Hydrogenation of the pterin derivatives over RANEY® Ni catalyst afforded dihydromonapterin and tetrahydromonapterin in optically active forms. We also investigated an alternative route involving an Amadori rearrangement, followed by the Polonovski-Boon reaction as the key step to make these monapterin derivatives.

In vitro retardation and modulation of human insulin amyloid fibrillation by Fe3+ and Cu2+ ions

Metal ions of Fe, Co, Ni, Cu, and Zn can form bonds through the carboxylate, hydroxyl, thiol, and imidazole side chains of proteins and those bonds are significantly more stable than those formed by non-transition metals.

Enhancing production and purity of 9-OH-AD from phytosterols by balancing metabolic flux of the side-chain degradation and 9-position hydroxylation in Mycobacterium neoaurum.

9α-Hydroxyandroster-4-ene-3,17-dione (9-OH-AD) is a representative steroid drug intermediate that can be prepared by phytosterols (PS) biotransformation with mycobacteria in a resting cell-cyclodextrin system. In this study, over-expression of 17β-hydroxysteroid dehydrogenase (Hsd4A)was testified to enhance the side-chain degradation of PS and to reduce the incomplete degradation by-products. Meanwhile, the complete degradation product 4-androstene-3,17-dione (AD)was increased due to the lack of 3-Ketosteroid 9α-Hydroxylase (KshA1) activities. To increase the production and purity of 9-OH-AD, the metabolic pathway of the side-chain degradation of PS and 9-position hydroxylationwas modulated by balancing the over-expression of Hsd4A and KshA1 in mycobacteria and reducing the bioconversion rate via lowering the ratio of PS and cyclodextrin. The production and purity of 9-OH-AD in brothwere improved from 22.18gL-1 and 77.13% to 28.27gL-1 and 87.84%, with a molar yield of 78.32%.

Microphase separation structures facilitated by dipole molecules and hydrogen bonding in poly (biphenyl piperidine-isatin) anion exchange membranes

The anion exchange membranes (AEMs) have the important function of blocking fuel and conducting OH− to make the cell form a circuit, so its alkali stability directly affects the performance of alkaline fuel cells. Currently, AEMs face a “trade-off” between ionic conductivity and stability. In order to solve this problem, we optimize the alkali resistance and conductivity of AEMs by introducing dipole-interacting side chains and hydrophilic hydroxyl side chains acting simultaneously on a rigid polymer backbone. By adopting the design strategy of “1 + 1>2″, the ion channel efficiency also makes a high-speed leap from a “two-wheeled bicycle” to a “four-wheeled car”. This effective microscopic modulation is clearly demonstrated by transmission electron microscope (TEM), where the AEMs after molecular design carry out a more obvious microphase separation structure at 200 nm. Through the PBP-54-IS- [OPD-100-OH-0] membrane, the maximum ionic conductivity is 100.62 mS cm−1 at 80 °C. After 1000 h at 80 °C, the retention rate of OH− conductivity of all AEMs was over 88.24 %. We also test its fuel cell performance for analysis. When the temperature of the two poles is 75 °C/72 °C, the power density is 147.28 mW cm−2. These are in line with the inherent requirements of AEMs for fuel cell applications.

Enzymatically Formed Oxysterols and Cell Death.

The side-chain hydroxylation of cholesterol by specific enzymes produces 24(S)-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol, and other products. These enzymatically formed side-chain oxysterols act as intermediates in the biosynthesis of bile acids and serve as signaling molecules that regulate cholesterol homeostasis. Besides these intracellular functions, an imbalance in oxysterol homeostasis is implicated in pathophysiology. Furthermore, growing evidence reveals that oxysterols affect cell proliferation and cause cell death. This chapter provides an overview of the pathophysiological role of side-chain oxysterols in developing human diseases. We also summarize our understanding of the molecular mechanisms underlying the induction of various forms of cell death by side-chain oxysterols.