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ACS Applied Materials & Interfaces Jun 2024We report herein the synthesis of three detergents bearing a perfluorinated cyclohexyl group connected through a short, hydrogenated spacer (i.e., propyl, butyl, or...
We report herein the synthesis of three detergents bearing a perfluorinated cyclohexyl group connected through a short, hydrogenated spacer (i.e., propyl, butyl, or pentyl) to a β-maltoside polar head that are, respectively, called FCymal-3, FCymal-4, and FCymal-5. Increasing the length of the spacer decreased the critical micellar concentration (CMC), as demonstrated by surface tension (SFT) and isothermal titration calorimetry (ITC), from 5 mM for FCymal-3 to 0.7 mM for FCymal-5. The morphology of the micelles was studied by dynamic light scattering (DLS), analytical ultracentrifugation (AUC), and small-angle X-ray scattering (SAXS), indicating heterogeneous rod-like shapes. While micelles of FCymal-3 and -4 have similar hydrodynamic diameters of ∼10 nm, those of FCymal-5 were twice as large. We also investigated the ability of the detergents to solubilize lipid membranes made of 1-palmitoyl-2-oleyl--glycero-3-phosphocholine (POPC). Molecular modeling indicated that the FCymal detergents generate disorder in lipid bilayers, with FCymal-3 being inserted more deeply into bilayers than FCymal-4 and -5. This was experimentally confirmed using POPC vesicles that were completely solubilized within 2 h with FCymal-3, whereas FCymal-5 required >8 h. A similar trend was noticed for the direct extraction of membrane proteins from membranes, with FCymal-3 being more potent than FCymal-5. An opposite trend was observed in terms of stabilization of the two model membrane proteins bacteriorhodopsin (bR) and SpNOX. In all three FCymal detergents, bR was stable for at least 2 months with no signs of aggregation. However, while the structural integrity of bR was fully preserved in FCymal-4 and -5, minor bleaching was observed in FCymal-3. Similarly, SpNOX exhibited the least activity in FCymal-3 and the highest activity in FCymal-5. By combining solubilizing and stabilizing potency, FCymal detergents push forward our expectations of the usefulness of fluorinated detergents for handling and investigating membrane proteins.
PubMed: 38885044
DOI: 10.1021/acsami.4c03359 -
Current Allergy and Asthma Reports Jun 2024Modernization and Westernization in industrialized and developing nations is associated with a substantial increase in chronic noncommunicable diseases. This... (Review)
Review
PURPOSE OF REVIEW
Modernization and Westernization in industrialized and developing nations is associated with a substantial increase in chronic noncommunicable diseases. This transformation has far-reaching effects on lifestyles, impacting areas such as economics, politics, social life, and culture, all of which, in turn, have diverse influences on public health. Loss of contact with nature, alternations in the microbiota, processed food consumption, exposure to environmental pollutants including chemicals, increased stress and decreased physical activity jointly result in increases in the frequency of inflammatory disorders including allergies and many autoimmune and neuropsychiatric diseases. This review aims to investigate the relationship between Western lifestyle and inflammatory disorders.
RECENT FINDINGS
Several hypotheses have been put forth trying to explain the observed increases in these diseases, such as 'Hygiene Hypothesis', 'Old Friends', and 'Biodiversity and Dysbiosis'. The recently introduced 'Epithelial Barrier Theory' incorporates these former hypotheses and suggests that toxic substances in cleaning agents, laundry and dishwasher detergents, shampoos, toothpastes, as well as microplastic, packaged food and air pollution damage the epithelium of our skin, lungs and gastrointestinal system. Epithelial barrier disruption leads to decreased biodiversity of the microbiome and the development of opportunistic pathogen colonization, which upon interaction with the immune system, initiates local and systemic inflammation. Gaining a deeper comprehension of the interplay between the environment, microbiome and the immune system provides the data to assist with legally regulating the usage of toxic substances, to enable nontoxic alternatives and to mitigate these environmental challenges essential for fostering a harmonious and healthy global environment.
PubMed: 38884832
DOI: 10.1007/s11882-024-01149-7 -
Journal of Structural Biology: X Jun 2024NMR spectroscopy has played a pivotal role in fragment-based drug discovery by coupling detection of weak ligand-target binding with structural mapping of the binding...
NMR spectroscopy has played a pivotal role in fragment-based drug discovery by coupling detection of weak ligand-target binding with structural mapping of the binding site. Fragment-based screening by NMR has been successfully applied to many soluble protein targets, but only to a limited number of membrane proteins, despite the fact that many drug targets are membrane proteins. This is partly because of difficulties preparing membrane proteins for NMR-especially human membrane proteins-and because of the inherent complexity associated with solution NMR spectroscopy on membrane protein samples, which require the inclusion of membrane-mimetic agents such as micelles, nanodiscs, or bicelles. Here, we developed a generalizable protocol for fragment-based screening of membrane proteins using NMR. We employed two human membrane protein targets, both in fully protonated detergent micelles: the single-pass C-terminal domain of the amyloid precursor protein, C99, and the tetraspan peripheral myelin protein 22 (PMP22). For both we determined the optimal NMR acquisition parameters, protein concentration, protein-to-micelle ratio, and upper limit to the concentration of D-DMSO in screening samples. Furthermore, we conducted preliminary screens of a plate-format molecular fragment mixture library using our optimized conditions and were able to identify hit compounds that selectively bound to the respective target proteins. It is hoped that the approaches presented here will be useful in complementing existing methods for discovering lead compounds that target membrane proteins.
PubMed: 38883400
DOI: 10.1016/j.yjsbx.2024.100100 -
3 Biotech Jul 2024Thermoalkaliphilic lipase enzymes are mostly favored for use in the detergent industry. While there has been considerable research on lipases, a significant portion of...
UNLABELLED
Thermoalkaliphilic lipase enzymes are mostly favored for use in the detergent industry. While there has been considerable research on lipases, a significant portion of these enzymes remains unexplored or undocumented in the scientific literature. This work performed in silico phylogeny, sequence alignment, structural and enzyme-substrate interaction analyses of the five thermoalkaliphilic lipases belonging to different species ( lipase = Lip, B4113_201601 lipase = Lip, HTA426 lipase = Lip, SP22 lipase = Lip, NTU 03 lipase = Lip). For this purpose, unreviewed enzyme sequences of five thermoalkaliphilic lipases were analyzed at sequence and phylogeny levels. 3D homology enzyme models were built, validated, and investigated by different bioinformatics tools. The ligand interactions screening using seven para-nitrophenyl (NP) esters and enzyme-ligand interactions were analyzed on Lip:NP-C12 and BTL2:NP-C12 by MD simulation. Biophysicochemical characteristic analysis showed that Lip had a theoretical value of above 65 ºC, and a higher aliphatic index indicating greater thermal stability. Sequence alignment showed a hydrophilic threonine in the α6 helix of Lip, indicating high enzymatic activity. A normalized temperature factor B (B'-factor) analysis showed that the lid domains of five lipases significantly possessed lower B'-factor values, compared to lipase 2 (BTL2), indicating that they had higher rigidity. Molecular docking results indicated that the five lipases had the highest binding affinity toward NP-C12. The RMSF investigation revealed that the thermostability of Lip is influenced by specific molecular elements: D202-S203 within the αB region of the lid domain, and E274-Q275 within the b3 strand, as well as W278 in the b3-b4 loop, and H282 in the b4 strand of the Ca-binding region. MD simulation analysis showed that catalytic residue S114 and at least one oxyanion hole residue (F17 and/or Q114) in Lip frequently formed hydrogen bonds with the NP-C12 ligand at 343 K and 348 K throughout the simulation process, indicating that Lip might catalyze relatively long-chain ligand NP-C12 with high performance. In conclusion, Lip might be a more suitable candidate as the detergent additive. In addition, this investigation can offer valuable perspectives on Family I.5 lipases such as Lip for future exploration in the field of protein engineering.
SUPPLEMENTARY INFORMATION
The online version contains supplementary material available at 10.1007/s13205-024-04023-5.
PubMed: 38882640
DOI: 10.1007/s13205-024-04023-5 -
ChemPlusChem Jun 2024Single particle cryo electron microscopy (cryo-EM) is now the major method for the determination of integral membrane protein structure. For the success of a given...
Single particle cryo electron microscopy (cryo-EM) is now the major method for the determination of integral membrane protein structure. For the success of a given project the type of membrane mimetic used for extraction from the native cell membrane, purification to homogeneity and finally cryo-grid vitrification is crucial. Although small molecule amphiphiles - detergents - are the most widely used membrane mimetic, specific tailoring of detergent structure for single particle cryo-EM is rare and the demand for effective detergents not satisfied. Here, we compare the popular detergent lauryl maltose-neopentyl glycol (LMNG) with the novel detergent neopentyl glycol-derived triglucoside-C11 (NDT-C11) in its behavior as free detergent and when bound to two types of multisubunit membrane protein complexes - cyanobacterial photosystem I (PSI) and mammalian F-ATP synthase. We conclude that NDT-C11 has high potential to become a very useful detergent for single particle cryo-EM of integral membrane proteins.
PubMed: 38881532
DOI: 10.1002/cplu.202400242 -
Yeast (Chichester, England) Jun 2024Cellobiose lipids are surface-active compounds or biological detergents produced by distinct Basidiomycetes yeasts, of which the most and best-described ones belong to...
Cellobiose lipids are surface-active compounds or biological detergents produced by distinct Basidiomycetes yeasts, of which the most and best-described ones belong to the Ustilaginomycetes class. The molecules display slight variation in congener type, which is linked to the hydroxylation position of the long fatty acid, acetylation profile of the cellobiose unit, and presence or absence of the short fatty acid. In general, this variation is strain specific. Although cellobiose lipid biosynthesis has been described for about 11 yeast species, hitherto only two types of biosynthetic gene clusters are identified, and this for only three species. This work adds six more biosynthetic gene clusters and describes for the first time a novel type of cellobiose lipid biosynthetic cluster with a simplified architecture related to specific cellobiose lipids synthesized by Trichosporonaceae family members.
PubMed: 38877753
DOI: 10.1002/yea.3969 -
BMC Plant Biology Jun 2024Selenium is essential for livestock and human health. The traditional way of adding selenium to livestock diets has limitations, and there is a growing trend to provide...
BACKGROUND
Selenium is essential for livestock and human health. The traditional way of adding selenium to livestock diets has limitations, and there is a growing trend to provide livestock with a safe and efficient source of selenium through selenium-enriched pasture. Therefore, this study was conducted to investigate the effects of selenium enrichment on fermentation characteristics, selenium content, selenium morphology, microbial community and in vitro digestion of silage alfalfa by using unenriched (CK) and selenium-enriched (Se) alfalfa as raw material for silage.
RESULTS
In this study, selenium enrichment significantly increased crude protein, soluble carbohydrate, total selenium, and organic selenium contents of alfalfa silage fresh and post-silage samples, and it significantly decreased neutral detergent fiber and acid detergent fiber contents (p < 0.05). Selenium enrichment altered the form of selenium in plants, mainly in the form of SeMet and SeMeCys, which were significantly higher than that of CK (p < 0.05). Selenium enrichment could significantly increase the lactic acid content, reduce the pH value, change the diversity of bacterial community, promote the growth of beneficial bacteria such as Lactiplantibacillus and inhibit the growth of harmful bacteria such as Pantoea, so as to improve the fermentation quality of silage. The in vitro digestibility of dry matter (IVDMD), in vitro digestibility of acid detergent fibers (IVADFD) and in vitro digestibility of acid detergent fibers (IVNDFD) of silage after selenium enrichment were significantly higher than those of CK (p < 0.05).
CONCLUSION
This study showed that the presence of selenium could regulate the structure of the alfalfa silage bacterial community and improve alfalfa silage fermentation quality. Selenium enrichment measures can change the morphology of selenium in alfalfa silage products, thus promoting the conversion of organic selenium.
Topics: Medicago sativa; Silage; Fermentation; Selenium; Microbiota; Animals; Animal Feed
PubMed: 38877393
DOI: 10.1186/s12870-024-05268-1 -
Archives of Biochemistry and Biophysics Jun 2024Mutation of phenylalanine at position 508 in the cystic fibrosis transmembrane conductance regulator (F508del CFTR) yields a protein unstable at physiological...
Mutation of phenylalanine at position 508 in the cystic fibrosis transmembrane conductance regulator (F508del CFTR) yields a protein unstable at physiological temperatures that is rapidly degraded in the cell. This mutation is present in about 90% of cystic fibrosis patients, hence there is great interest in compounds reversing its instability. We have previously reported the expression of the mutated protein at low temperature and its purification in detergent. Here we describe the use of the protein to screen compounds present in a library of Federal Drug Administration (FDA) - approved drugs and also in a small natural product library. The kinetics of unfolding of F508del CFTR at 37 °C were probed by the increase in solvent-exposed cysteine residues accessible to a fluorescent reporter molecule. This occurred in a bi-exponential manner with a major (≈60%) component of half-life around 5 min and a minor component of around 60 min. The faster kinetics match those observed for loss of channel activity of F508del CFTR in cells at 37 °C. Most compounds tested had no effect on the fluorescence increase, but some were identified that significantly slowed the kinetics. The general properties of these compounds, and any likely mechanisms for inducing stability in purified CFTR are discussed. These experimental data may be useful for artificial intelligence - aided design of CFTR-specific drugs and in the identification of stabilizing additives for membrane proteins (in general).
PubMed: 38876247
DOI: 10.1016/j.abb.2024.110050 -
Biofabrication Jun 2024Recent advancements in 3D cancer modeling have significantly enhanced our ability to delve into the intricacies of carcinogenesis. Despite the pharmaceutical industry's...
Recent advancements in 3D cancer modeling have significantly enhanced our ability to delve into the intricacies of carcinogenesis. Despite the pharmaceutical industry's substantial investment of both capital and time in the drug screening and development pipeline, a concerning trend persists: drug candidates screened on conventional cancer models exhibit a dismal success rate in clinical trials. One pivotal factor contributing to this discrepancy is the absence of drug testing on pathophysiologically biomimetic 3D cancer models during pre-clinical stages. Unfortunately, current manual methods of 3D cancer modeling, such as spheroids and organoids, suffer from limitations in reproducibility and scalability. In our study, we have meticulously developed 3D bioprinted breast cancer model utilizing decellularized adipose tissue-based hydrogel obtained via a detergent-free decellularization method. Our innovative printing techniques allows for rapid, high-throughput fabrication of 3D cancer models in a 96-well plate format, demonstrating unmatched scalability and reproducibility. Moreover, we have conducted extensive validation, showcasing the efficacy of our platform through drug screening assays involving two potent anti-cancer drugs, 5-Fluorouracil and PRIMA-1. Notably, our platform facilitates effortless imaging and gene expression analysis, streamlining the evaluation process. In a bid to enhance the relevance of our cancer model, we have introduced a heterogeneous cell population into the DAT-based bioink. Through meticulous optimization and characterization, we have successfully developed a biomimetic immunocompetent breast cancer model, complete with microenvironmental cues and diverse cell populations. This breakthrough paves the way for rapid multiplex drug screening and the development of personalized cancer models, marking a paradigm shift in cancer research and pharmaceutical development.
Topics: Humans; Breast Neoplasms; Female; Bioprinting; High-Throughput Screening Assays; Printing, Three-Dimensional; Drug Screening Assays, Antitumor; Cell Line, Tumor; Drug Evaluation, Preclinical; Hydrogels; Antineoplastic Agents; Tissue Engineering; Adipose Tissue; Models, Biological; Reproducibility of Results
PubMed: 38876096
DOI: 10.1088/1758-5090/ad586b -
Phytopathology Jun 2024The Fusarium head blight (FHB) pathogen Fusarium graminearum produces the trichothecene mycotoxin deoxynivalenol (DON) and reduces wheat yield and grain quality. Spring...
The Fusarium head blight (FHB) pathogen Fusarium graminearum produces the trichothecene mycotoxin deoxynivalenol (DON) and reduces wheat yield and grain quality. Spring wheat (Triticum aestivum L.) genotype CB037 was transformed with constitutive expression (CE) constructs containing sorghum (Sorghum bicolor L. (Moench)) genes encoding monolignol biosynthetic enzymes, caffeoyl-Coenzyme A (CoA) 3-O-methyltransferase (SbCCoAOMT), 4-coumarate-CoA ligase (Sb4CL), or coumaroyl shikimate 3-hydroxylase (SbC3'H), or monolignol pathway transcriptional activator, SbMyb60. Spring wheats were screened for Type I (resistance to initial infection, using spray inoculations) and Type II (resistance to spread within the spike, using single floret inoculations) resistances in the field (spray) and greenhouse (spray and single floret). Following field inoculations, disease index, percent Fusarium damaged kernels (FDK), and DON measurements of CE plants were similar to or greater than CB037. For greenhouse inoculations, the area under the disease progress curve (AUDPC) and FDK were determined. Following screens, focus was placed on two each, SbC3'H and SbCCoAOMT CE lines because of trends towards decreased AUDPC and FDK observed following single floret inoculations. These four lines were as susceptible as CB037 following spray inoculations. However, single floret inoculations showed that these CE lines had significantly reduced AUDPC (P<0.01) and FDK (P≤0.02) compared with CB037, indicating improved Type II resistance. None of these CE lines had increased acid detergent lignin, as compared with CB037, indicating that lignin concentration may not be a major factor in FHB resistance. The SbC3'H and SbCCoAOMT CE lines are valuable for investigating phenylpropanoid-based resistance to FHB.
PubMed: 38875177
DOI: 10.1094/PHYTO-01-24-0005-R