Binding Process Research Articles

Natural compounds inhibit Monkeypox virus methyltransferase VP39 in silico studies

VP39, an essential 2′-O-RNA methyltransferase enzyme discovered in Monkeypox virus (MPXV), plays a vital role in viral RNA replication and transcription. Inhibition of the enzyme may prevent viral replication. In this context, using a combination of molecular docking and molecular dynamics (MDs) simulations, the inhibitory ability of NCI Diversity Set VII natural compounds to VP39 protein was investigated. It should be noted that the computed binding free energy of ligand via molecular docking and linear interaction energy (LIE) approaches are in good agreement with the corresponding experiments with coefficients of R = 0.72 and 0.75, respectively. NSC 319990, NSC 196515 and NSC 376254 compounds were demonstrated that can inhibit MPVX methyltransferase VP39 protein with the similar affinity compared to available inhibitor sinefungin. Moreover, nine residues involving Gln39, Gly68, Gly72, Asp95, Arg97, Val116, Asp138, Arg140 and Asn156 may be argued that they play an important role in binding process of inhibitors to VP39. Communicated by Ramaswamy H. Sarma

Unraveling Bacterial Single-Stranded Sequence Specificities: Insights from Molecular Dynamics and MMPBSA Analysis of Oligonucleotide Probes.

We utilized molecular dynamics (MD) simulations and Molecular Mechanics Poisson-Boltzmann Surface Area (MMPBSA) free energy calculations to investigate the specificity of two oligonucleotide probes, namely probe B and probe D, in detecting single-stranded DNA (ssDNA) within three bacteria families: Enterobacteriaceae, Pasteurellaceae, and Vibrionaceae. Due to the limited understanding of molecular mechanisms in the previous research, we have extended the discussion to focus specifically on investigating the binding process of bacteria-probe DNA duplexes, with an emphasis on analyzing the binding free energy. The role of electrostatic contributions in the specificity between the oligonucleotide probes and the bacterial ssDNAs was investigated and found to be crucial. Our calculations yielded results that were highly consistent with the experimental data. Through our study, we have successfully exhibited the benefits of utilizing in-silico approaches as a powerful virtual-screening tool, particularly in research areas that demand a thorough comprehension of molecular interactions.

Biophysical and structural characterization of tetramethrin serum protein complex and its toxicological implications.

Tetramethrin (TMT) is a commonly used insecticide and has a carcinogenic and neurodegenerative effect on humans. The binding mechanism and toxicological implications of TMT to human serum albumin (HSA) were examined in this study employing a combination of biophysical and computational methods indicating moderate binding affinity and potential hepato and renal toxicity. Fluorescence quenching experiments showed that TMT binds to HSA with a moderate affinity, and the binding process was spontaneous and predominantly enthalpy-driven. Circular dichroism spectroscopy revealed that TMT binding did not induce any significant conformational changes in HSA, resulting in no changes in its alpha-helix content. The binding site and modalities of TMT interactions with HSA as computed by molecular docking and molecular dynamics simulations revealed that it binds to Sudlow site II of HSA via hydrophobic interactions through its dimethylcyclopropane carboxylate methyl propanyl group. The structural dynamics of TMT induce proper fit into the binding site creating increased and stabilizing interactions. Additionally, molecular mechanics-Poisson Boltzmann surface area calculations also indicated that non-polar and van der Waals were found to be the major contributors to the high binding free energy of the complex. Quantum mechanics (QM) revealed the conformational energies of the binding confirmation and the degree of deviation from the global minimum energy conformation of TMT. The results of this study provide a comprehensive understanding of the binding mechanism of TMT with HSA, which is important for evaluating the toxicity of this insecticide in humans.

Revealing the binding mechanism of BACE1 inhibitors through molecular dynamics simulations

Alzheimer’s disease is a debilitating neurodegenerative disorder, and the Beta-Site Amyloid Precursor Protein Cleaving Enzyme 1 (BACE1) is a key therapeutic target in its treatment. This study employs molecular dynamics simulations and binding energy analysis to investigate the binding interactions between BACE1 and four selected small molecules: CNP520, D9W, NB641, and NB360. The binding model analysis indicates that the binding of BACE1 with four molecules are stable, except the loop regions show significant fluctuation. The binding free energy analyses reveal that NB360 exhibits the highest binding affinity with BACE1, surpassing other molecules (CNP520, D9W, and NB641). Detailed energy component assessments highlight the critical roles of electrostatic interactions and van der Waals forces in the binding process. Furthermore, residue contribution analysis identifies key amino acids influencing the binding process across all systems. Hydrogen bond analysis reveals a limited number of bonds between BACE1 and each small molecule, highlighting the importance of structural modifications to enable more stable hydrogen bonds. This research provides valuable insights into the molecular mechanisms of potential Alzheimer’s disease therapeutics, guiding the way for improved drug design and the development of effective treatments targeting BACE1. Communicated by Ramaswamy H. Sarma

Assessment of Phytotoxicity of Potable Water Treatment Plant Sludge-bound Compost Pellets on Seed Germination of Radish (Raphanus sativus L.)

Binding loose compost into pellets needs different binding materials. Maintenance of pellet stability highly depends on the binding materials used. Potable water treatment plant sludge (WTPS) has the potential to be used as a binding material due to the high content of clay, organic matter, and nutrients. However, compost and WTPS may contain heavy metals, toxic compounds, salts, and growth inhibitors. Therefore, the determination of the phytotoxicity of WTPS bound compost pellets is essential before amending them to the soil. The present study aimed to assess the phytotoxicity of different pelleted compost using seed germination bioassay of Raphanus sativus L. Four compost pellets were considered (T1: commercial compost pellet (100% compost), T2: commercial integrated pellet (90% compost+10% inorganic fertilizer), T3: WTPS-bound compost pellet (90% compost+10% WTPS), T4: WTPS-bound integrated pellet (80% compost+10% WTPS+10% inorganic fertilizer)) and pellet aqueous extracts (PAE) were prepared, respectively. Distilled water was used as the control. The dilution sequence of PAE as 50% and 100% were tested for seed germination in Petri dishes in a randomized design with three replicates. pH, EC, and selected heavy metals (Al, Zn, Cu and Cr) were determined in PAE. RSG% (Relative Seed Germination %), RRG% (Relative radicle Growth %), and GI% (Germination Index) were calculated for all the PAE after 72 hours. The PAE had a pH range from 6.8-7.2. The Cr was not detected in all 100% PEA. The highest levels of Al and Zn were detected in 100% PAE of T1. All the treatments in both 50% and 100% PAE showed GI% higher than 80% except the 100% PAE of commercial integrated pellet (T2) and WTPS-bound integrated pellet (T4). The PAE from the T2 and T4 showed low RSG% and RRG% and thereby the GI% is low due to high EC in PAE. The GI% of 100% PAE in T2 and T4 was 32.33% and 67.25%, respectively. The lowest values for RSG% and RRG% were recorded for T2 as 44.44% and 72.75%, respectively. Spearman‘s Rank Correlation coefficients were calculated for EC with RSG%, RRG%, and GI% and all variables showed negative correlations as (-0.44), (-0.97), and (-0.69), respectively. This indicates that high EC reduces radical growth and seed germination. PAE at 100% concentration levels of commercial integrated pellet showed high phytotoxicity, and WTPS-bound integrated pellet showed less phytotoxicity while other treatments did not show any sign of phytotoxicity. All the PAE at 50% concentration level were free from phytotoxicity. Hence, 10% WTPS in w/w basis can be used as a binding agent in pelletizing loose compost. Different WTPS concentrations should be tested in binding process for its optimum utilization. 
 Keywords: Bioassay, Compost, Phytotoxicity, Radish, Water treatment plant sludge

Integrating Label-free proteomics and molecular dynamics to investigate the interactions between proteins and bioactive compounds in Rosa roxburghii tratt fermented by Lactobacillus acidophilus GIM1.208

Lactobacillus acidophilus GIM1.208 (LA) can improve flavor, enhance antioxidant and hypoglycemic activities of Rosa roxburghii Tratt (RRT). This study investigated the interactions between proteins and active compounds in RRT fermented by LA to reveal the underlying reasons for changes in flavor and activity. A total of 6239 proteins were characterized and quantified by Label-free proteomics, with their molecular weights were mainly concentrated in the range of 10–40 kDa. The 129 differential proteins screened were mainly enriched in compounds synthesis or transformation pathways, such as glycolysis. Integrated analysis of proteomics and metabolomics revealed a strong correlation between proteins and metabolites, especially β-glucosidase was closely correlation with flavonoids and triterpenoids. Additionally, molecular dynamics (MD) simulations have demonstrated that glycosides establish hydrogen bonds with the amino acids of β-glucosidase, while hydrophobic interactions serve as the primary driving force behind the binding process. Notably, Rosamutin exhibited the most significant binding energy with β-glucosidase (−66.257 kJ/mol). The results of the MD analysis further demonstrate the huge potential of LA β-glucosidase to transform glycosides in RRT. In summary, proteins in LA fermented RRT contribute to the consumption or synthesis of compounds, ultimately enhancing the flavor and activity of RRT. This study's results can provide a reference for the research of proteins and bioactive compounds of RRT.

Nanocomplex of (−)-epigallocatechin-3-gallate and glycated ovalbumin based on natural deep eutectic solvents: Binding analysis and functional improvement

Natural deep eutectic solvents (NADES) are considered an efficient reaction medium for the production of glycated proteins. By optimizing the Maillard reaction conditions through response surface analysis, a glycation product (GOVA) of ovalbumin (OVA) with high grafting degree (92.2%) was prepared in choline chloride/glucose NADES. The binding information of (−)-epigallocatechin-3-gallate (EGCG) to OVA/GOVA was acquired by fluorescence spectroscopy and molecular docking. The interaction strength of GOVA + EGCG was higher than that of OVA + EGCG. Hydrogen bonding and van der Waals forces were the major driving forces in the binding process. Glycation and EGCG binding reduced the surface hydrophobicity, α-helix content and digestibility of OVA. Compared with OVA, the particle sizes of GOVA and GOVA + EGCG were increased by 54.8% and 87.7%, respectively. In addition, glycation increased the emulsifying activity index and foaming capacity of OVA by 32.6% and 20.0%, respectively, which was further increased in the presence of EGCG. The encapsulation of OVA, especially GOVA, improved the antioxidant activity, stability and bioaccessibility of EGCG. The results suggest that the glycated proteins based on NADES may provide an effective alternative for the protection and transport of bioactive ingredients in the food industries.

Molecular mechanism of NAD+ and NMN binding to the Nudix homology domains of DBC1

Deleted in breast cancer 1 (DBC1) is a human nuclear protein that modulates the activities of various proteins involved in cell survival and cancer progression. Oxidized form of nicotinamide adenine dinucleotide (NAD+) is suggested to bind to the Nudix homology domains (NHDs) of DBC1, thereby regulating DBC1-Poly (ADP-ribose) polymerase 1 (PARP1) interactions, resulting in the restoration of DNA repair. Using Nuclear Magnetic Resonance (NMR) and Isothermal Titration Calorimetry (ITC), we confirmed NAD+ and its precursor nicotinamide mononucleotide (NMN) both bind the NHD domain of DBC1 (DBC1354–396). NAD+ likely interacts with DBC1354–396 through hydrogen bonding, with a binding affinity (8.99 μM) nearly twice that of NMN (17.0 μM), and the key binding sites are primarily residues E363 and D372, in the agreement with Molecular Docking experiments. Molecular Dynamics (MD) simulation further demonstrated E363 and D372's anchoring role in the binding process. Additional mutagenesis experiments of E363 and D372 confirmed their critical involvement of ligand-protein interactions. These findings lead to a better understanding of how NAD+ and NMN regulate DBC1, thereby offering insights for the development of targeted therapies and drug research focused on DBC1-associated tumors.

Interaction between curcumin and ultrafiltered casein micelles or whey protein, and characteristics of their complexes.

This work evaluated the interaction between micellar casein (MC) or whey protein (WP) in ultrafiltration retentate with curcumin (Cur), as well as the physicochemical and functional properties of Cur-MC and Cur-WP complexes. The MC had a higher affinity for Cur than WP, shown by higher binding constants of Cur-MC at various temperatures. Thermodynamic analysis of the binding process indicated that the interaction between Cur and MC or WP was hydrophobic in nature. Cur promoted the size and polydispersity index of MC and WP at 4mM but did not alter the morphology of spray-dried MC and WP. The Cur-MC complexes showed better aqueous solubility at pH 2-3 and 6-10 compared to free MC. Combination with MC or WP improved the 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonate) radical scavenging activity of Cur. In addition, combination with MC and WP promoted cumulative release of Cur during simulated gastrointestinal digestion, especially for WP. Thus, MC and WP in retentates can be good alternative protein-based carriers for Cur delivery, whereas their complexes in powder form have good functional properties that could be used as active food ingredients in several food formulations. PRACTICAL APPLICATION: Microfiltration is a cheap and convenient approach that can be used to easily produce micellar casein (MC), with whey protein (WP) as one byproduct. In this study, we proved that MC and WP in retentates have strong interaction with curcumin (Cur), whereas their complexes have good functional properties. Thus, spray-dried MC-Cur or WP-Cur complexes could be used as active food ingredients in several food formulations.

Spectroscopic insights with molecular docking and molecular dynamic simulation studies of anticancer drug 5-fluororouracil targeting human pyruvate kinase m2

This study was conducted to test the efficacy of 5-fluorouracil (5-FU) as an anticancer drug against the human pyruvate kinase isozyme M2 (PKM2) using spectroscopic, molecular docking and molecular dynamic simulation studies. PKM2 fluorescence quenching studies in the presence of 5-FU performed at three different temperatures indicates dynamic quenching processes with single-set of binding (n ≈ 1) profile. The biomolecular quenching constants (k q) and the effective binding constants (K b) obtained are shown to increase with temperature. The calculated enthalpy (ΔH) and entropy changes (ΔS) are estimated to be −118.06 kJ/mol and 146.14 kJ/mol/K respectively, which suggest the possible mode of interaction as electrostatic and hydrogen bonding. Further, these values were used to estimate the free energy changes (ΔG) and that increases with temperature. The negative ΔG values clearly indicates spontaneous binding process that stabilizes the complex formed between 5-FU and PKM2. Far-UV CD spectra of PKM2 in the presence of 5-FU shows decrease in α-helix contents which point towards the destabilization of secondary structure that weakens the biological activity of PKM2. The intrinsic fluorescence study and circular dichroism (CD) spectra showed minor conformational changes of PKM2 in the presence of 5-FU. Additionally, the results obtained from molecular docking and all-atom molecular dynamic simulation study supports the insight of the spectroscopic binding studies, and strengthens the dynamic stability of the complex between 5-FU and PKM2 through H-bonding. This study establishes a paradigm of 5-FU-PKM2 complexation and the efficacy of 5-FU that compromises the biological activity of the targeted PKM2. Communicated by Ramaswamy H. Sarma

Exploring the mechanism of interaction between TBG and halogenated thiophenols: Insights from fluorescence analysis and molecular simulation

Thyroxine-binding globulin (TBG) plays a vital role in regulating metabolism, growth, organ differentiation, and energy homeostasis, exerting significant effects in various key metabolic pathways. Halogenated thiophenols (HTPs) exhibit high toxicity and harmfulness to organisms, and numerous studies have demonstrated their thyroid-disrupting effects. To understand the mechanism of action of HTPs on TBG, a combination of competitive binding experiments, multiple fluorescence spectroscopy techniques, molecular docking, and molecular simulations was employed to investigate the binding mechanism and identify the binding site. The competition binding assay between HTPs and ANS confirmed the competition of HTPs with thyroid hormone T4 for the active site of TBG, resulting in changes in the TBG microenvironment upon the binding of HTPs to the active site. Key amino acid residues involved in the binding process of HTPs and TBG were further investigated through residue energy decomposition. The distribution of high-energy contributing residues was determined. Analysis of root-mean-square deviation (RMSD) demonstrated the stability of the HTPs-TBG complex. These findings confirm the toxic mechanism of HTPs in thyroid disruption, providing a fundamental reference for accurately assessing the ecological risk of pollutants and human health. Providing mechanistic insights into how HTPS causes thyroid diseases.

Ice Recrystallization Unveils the Binding Mechanism Operating at a Diffused Interface.

Recrystallization of ice is a natural phenomenon that causes adverse effects in cryopreservation, agriculture, and in frozen food industry. It has long been recognized that ice recrystallization occurs through the Ostwald ripening and accretion processes. However, neither of these processes has been explored in microscopic detail by state-of-the-art experimental techniques. We carried out atomistic molecular dynamics (MD) simulations to explore ice recrystallization through the accretion process. Attempts have been made to elucidate the binding mechanism that is operating at the diffused ice-water interface. It is demonstrated that two ice crystals spontaneously recognize each other and bind together to form a large crystal in liquid water, resulting in ice recrystallization by accretion. Interestingly, the study reveals that the binding occurs due to the freezing of the interfacial water layer present between the two ice planes, even at a temperature above the melting point of the ice crystal. The synergistically enhanced ordering effect of two ice surfaces on the interfacial water leads to such freezing occurring during the binding process. However, proper crystallographic alignment is not necessarily required for the binding of the two crystals. Simulations have also been carried out to study the binding between an ice crystal and the model ice-binding surface (IBS) of an antifreeze protein above the melting point of the ice crystal. It is found that such binding at the IBS is accompanied by freezing of the interfacial water. This establishes that the synergetic ordering-driven freezing of interfacial water is a common binding mechanism at the diffused surfaces of ice crystals. We believe that this mechanism will provide a microscopic understanding of the process of recrystallization inhibition and thus help in designing suitable materials for potent applications in recrystallization inhibition.

Distinct mechanisms underlying the therapeutic effects of low-molecular-weight heparin and chondroitin sulfate on Parkinson's disease

Parkinson's disease (PD) is a neurodegenerative disorder influenced by various factors, including age, genetics, and the environment. Current treatments provide symptomatic relief without impeding disease progression. Previous studies have demonstrated the therapeutic potential of exogenous heparin and chondroitin sulfate in PD. However, their therapeutic mechanisms and structure-activity relationships remain poorly understood. In this study, low-molecular-weight heparin (L-HP) and chondroitin sulfate (L-CS) exhibited favorable therapeutic effects in a mouse model of PD. Proteomics revealed that L-HP attenuated mitochondrial dysfunction through its antioxidant properties, whereas L-CS suppressed neuroinflammation by inhibiting platelet activation. Two glycosaminoglycan (GAG)-binding proteins, manganese superoxide dismutase (MnSOD2) and fibrinogen beta chain (FGB), were identified as potential targets of L-HP and L-CS, and we investigated their structure-activity relationships. The IdoA2S-GlcNS6S/GlcNAc6S unit in HP bound to SOD2, whereas the GlcA-GalNAc4S and GlcA-GalNAc4S6S units in CS preferred FGB. Furthermore, N-S and 2-O-S in L-HP, and 4-O-S, 6-O-S, and -COOH in L-CS contributed significantly to the binding process. These findings provide new insights and evidence for the development and use of glycosaminoglycan-based therapeutics for PD.

Inhibitory mechanism of chrysin and diosmetin to α-glucosidase: insights from kinetics, multispectroscopy and molecular docking investigations

Inhibition of α-glucosidase activity is a promising method to prevent postprandial hyperglycemia. The inhibitory effect and interaction of chrysin and diosmetin on α-glucosidase were studied in this study. The results of inhibition kinetics showed that chrysin and diosmetin reversibly inhibited α-glucosidase activity with IC50 value of 26.445 ± 1.406 μmol L−1 and 18.380 ± 1.264 μmol L−1, respectively. Further research revealed that chrysin exhibited a mixed-type inhibitory pattern against α-glucosidase, while diosmetin was noncompetitive inhibitory with Ki value of (2.6 ± 0.04) ×10−4 mol L−1. Fluorescence spectroscopy showed that both chrysin and diosmetin could quench the intrinsic fluorescence of α-glucosidase, the maximum emission wavelength of tyrosine (Tyr) and tryptophan (Trp) were not moved by chrysin, but red shifted by diosmetin. UV-Vis, fourier transform infrared spectroscopy (FT-IR) and circular dichroism (CD) measurements showed that the secondary structure and microenvironment of α-glucosidase were changed by chrysin and diosmetin. Further analysis of molecular docking showed that chrysin and diosmetin could bind with α-glucosidase and might cause the decrease of α-glucosidase activity. The results of molecular dynamics (MD) simulation showed that the stability of chrysin (or diosmetin)-α-glucosidase complex system was changed during binding process. In conclusion, chrysin and diosmetin are good α-glucosidase inhibitors. Communicated by Ramaswamy H. Sarma

Revealing the Antiperspirant Components of Floating Wheat and Their Mechanisms of Action through Metabolomics and Network Pharmacology.

Floating wheat is a classical herbal with potential efficacy in the treatment of hyperhidrosis. Aiming at revealing the main components and potential mechanisms of floating wheat, a comprehensive and unique phytopharmacology profile study was carried out. First, common wheat was used as a control to look for chemical markers of floating wheat. In the screening analysis, a total of 180 shared compounds were characterized in common wheat and floating wheat, respectively. The results showed that floating wheat and common wheat contain similar types of compounds. In addition, in non-targeted metabolomic analysis, when taking the contents of the constituents into account, it was found that there indeed existed quite a difference between floating wheat and common wheat and 17 potential biomarkers for floating wheat. Meanwhile, a total of seven components targeted for hyperhidrosis were screened out based on network pharmacology. Seven key differential components were screened, among which kaempferol, asiatic acid, sclareol, enoxolone, and secoisolariciresinol had higher degree values than the others. The analysis of interacting genes revealed three key genes, namely, MAP2K1, ESR1, and ESR2. The Kyoto Encyclopaedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses showed that various signaling pathways were involved. Prolactin signaling, thyroid cancer, endocrine resistance, gonadotropin secretion, and estrogen signaling pathways were the main pathways of the intervention of floating wheat in excessive sweating, which was associated with the estrogenic response, hormone receptor binding, androgen metabolism, apoptosis, cancer, and many other biological processes. Molecular docking showed that the screened key components could form good bindings with the target proteins through intermolecular forces. This study reveals the active ingredients and potential molecular mechanism of floating wheat in the treatment of hyperhidrosis and provides a reference for subsequent basic research.

A chondroitin sulfate purified from shark cartilage and bovine serum albumin interaction activity

Chondroitin sulfate (CS) was extracted and purified from shark cartilage, and its interaction with bovine serum albumin (BSA) were studied. The content of chondroitin sulfate in shark cartilage was 29.97 % using the 1,9-dimethyl-methylene blue method. The molecular weight of CS was determined to be 62.464 kDa by high-performance gel permeation chromatography. UV and FT-IR spectroscopy identified the characteristics of CS and its functional group information. NMR spectroscopy and disaccharide derivatization revealed that CS was predominantly composed of disulfated disaccharides, specifically ΔDi4,6S. Fluorescence quenching experiments indicated that the interaction between CS and BSA exhibited static quenching, with a binding site number of 1. The binding process was primarily mediated by van der Waals forces and hydrogen bonds. Furthermore, synchronous and 3D fluorescence spectroscopy demonstrated that CS had minimal impact on the polarity and hydrophobicity of the microenvironment surrounding Tyr and Trp residues. UV–vis absorption and circular dichroism (CD) spectroscopy demonstrated the altered structure of BSA. The molecular docking analysis revealed that CS formed hydrogen bonds and salt bridges with BSA, predominantly binding to the IIA substructure domain of BSA. Investigating the interaction between CS and BSA holds the potential for enhancing its applications in drug delivery and tissue engineering endeavors.

Selective capture of uranium by p-block bismuth-based metal–organic framework

Energy shortage severely impedes the advancement of human society and economy. The vigorous development of nuclear energy is a viable solution to address this issue. The escalating depletion of uranium resources necessitates prioritizing the selective extraction of uranyl from seawater and wastewater to ensure the prosperity of nuclear power. A bismuth-based MOF, CAU-17, was synthesized by the one-step solvothermal method for targeted selection of uranyl in the first time. The uranium adsorption tests demonstrated that CAU-17 exhibited a high adsorption capacity of 421.9 mg g−1 (T = 25 °C, pH = 7), exceptional selectivity, outstanding salt tolerance, and favorable stability with 83.6 % of the initial capacity retained after 5 cycles. Through the utilization of XPS and theoretical calculations, it was elucidated that the carboxyl oxygen that connects the Bi played a pivotal role in the binding process between CAU-17 and uranium ions, with bismuth acting as an electron regulator through its bridging effect with carboxyl oxygen. This work not only presents an exceptional uranium adsorption material but also elucidates the role of core metal atoms in MOFs during the binding to uranium process, thereby offering a novel perspective for future advancements in MOFs-based adsorbents.

Stereoselective recognition of morphine enantiomers by μ-opioid receptor.

Stereospecific recognition of chiral molecules plays a crucial role in biological systems. The μ-opioid receptor (MOR) exhibits binding affinity towards (-)-morphine, a well-established gold standard in pain management, while it shows minimal binding affinity for the (+)-morphine enantiomer, resulting in a lack of analgesic activity. Understanding how MOR stereoselectively recognizes morphine enantiomers has remained a puzzle in neuroscience and pharmacology for over half-a-century due to the lack of direct observation techniques. To unravel this mystery, we constructed the binding and unbinding processes of morphine enantiomers with MOR via molecular dynamics simulations to investigate the thermodynamics and kinetics governing MOR's stereoselective recognition of morphine enantiomers. Our findings reveal that the binding of (-)-morphine stabilizes MOR in its activated state, exhibiting a deep energy well and a prolonged residence time. In contrast, (+)-morphine fails to sustain the activation state of MOR. Furthermore, the results suggest that specific residues, namely D1142.50 and D1473.32, are deprotonated in the active state of MOR bound to (-)-morphine. This work highlights that the selectivity in molecular recognition goes beyond binding affinities, extending into the realm of residence time.

A novel model for predicting prognosis in patients with idiopathic pulmonary fibrosis based on endoplasmic reticulum stress-related genes.

Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic disease of unknown pathogenic origin. Endoplasmic reticulum (ER) stress refers to the process by which cells take measures to ER function when the morphology and function of the reticulum are changed. Recent studies have demonstrated that the ER was involved in the evolution and progression of IPF. In this study, we obtained transcriptome data and relevant clinical information from the Gene Expression Omnibus database and conducted bioinformatics analysis. Among the 544 ER stress-related genes (ERSRGs), 78 were identified as differentially expressed genes (DEGs). These DEGs were primarily enriched in response to ER stress, protein binding, and protein processing. Two genes (HTRA2 and KTN1) were included for constructing an accurate molecular signature. The overall survival of patients was remarkably worse in the high-risk group than in the low-risk group. We further analyzed the difference in immune cells between high-risk and low-risk groups. M0 and M2 macrophages were significantly increased in the high-risk group. Our results suggested that ERSRGs might play a critical role in the development of IPF by regulating the immune microenvironment in the lungs, which provide new insights on predicting the prognosis of patients with IPF.

Novel Polystyrene-Binding Nanobody for Enhancing Immunoassays: Insights into Affinity, Immobilization, and Application Potential.

Nanobodies, which represent the next generation of antibodies due to their unique properties, face a significant limitation in their poor physical adsorption on solid supports. In this study, we successfully discovered polystyrene binding nanobodies from a synthetic nanobody library. Notably, bivalent nanobody B2 exhibited high affinity for polystyrene (0.7 nM for ELISA saturation binding analysis and 15.6 nM for isothermal titration calorimetry), displaying a pH-dependent behavior. Remarkably, hydrophobic and electrostatic interactions contribute minimally to the binding process. Molecular modeling provided insights into the interaction between B2 and polystyrene, revealing that the Trp51 residue within the CDR2 loop formed an aromatic H-bond with polystyrene at a distance of 2.74 Å, thus explaining the observed reduction in B2 affinity caused by Trp51 mutations. To explore B2's potential in protein immobilization, we constructed a bispecific nanobody by fusing B2 to an anticarcinoembryonic antigen nanobody 11C12, which cannot be immobilized on polystyrene through passive adsorption. Remarkably, the fusion construct achieved effective immobilization on polystyrene within 5 min by passing the need for periplasmic protein purification despite its low expression level. Moreover, the fusion construct demonstrated excellent linearity in the chemiluminescent enzyme immunoassay. For the first time, this study reports a simplified and seamless platform for the oriented immobilization of nanobody. Importantly, the entire process eliminated the need for protein purification, enabling efficient and rapid immobilization of fusion proteins directly from crude cell extracts, even when the expression level was low. Our developed process dramatically reduced the processing time from 2.5 days to just 5 min.