Β-pleated Sheet Research Articles

Harnessing ultrasonic power to optimize quinoa byproduct protein for sustainable utilization

The objective of this study was to elucidate the impact of ultrasonic treatment on quinoa by-product protein (QBP). Ultrasound treatment resulted in decreases in the moisture, lipid content, water activity, bulk and compact densities, and turbidity of the QBP. This treatment disrupted the compact protein structure and hydrophobic regions, thereby releasing a greater quantity of amino acids. Chemical analysis revealed that subjecting QBP aggregates to 400 W ultrasound transformed them into smaller and more uniformly distributed aggregates. Ultrasound significantly affected the secondary structure of QBP, increasing the relative proportions of α-helix and β-turn, while reducing β-pleated sheet and random coil proportions. Additionally, it altered the intrinsic fluorescence intensity and increased the sulfhydryl-group content. Notably, at 400 W, the protein exhibited optimal solubility, foaming properties, water and oil-binding capacity, and in vitro digestibility. These findings suggest that ultrasonic treatment can effectively enhance the quality and functional characteristics of quinoa protein byproducts, offering new perspectives for the comprehensive utilization of quinoa waste.

Amyloid Neuropathy: From Pathophysiology to Treatment in Light-Chain Amyloidosis and Hereditary Transthyretin Amyloidosis.

Amyloid neuropathy is caused by deposition of insoluble β-pleated amyloid sheets in the peripheral nervous system. It is most common in: (1) light-chain amyloidosis, a clonal non-proliferative plasma cell disorder in which fragments of immunoglobulin, light or heavy chain, deposit in tissues, and (2) hereditary transthyretin (ATTRv) amyloidosis, a disorder caused by autosomal dominant mutations in the TTR gene resulting in mutated protein that has a higher tendency to misfold. Amyloid fibrils deposit in the endoneurium of peripheral nerves, often extensive in the dorsal root ganglia and sympathetic ganglia, leading to atrophy of Schwann cells in proximity to amyloid fibrils and blood-nerve barrier disruption. Clinically, amyloid neuropathy is manifested as a length-dependent sensory predominant neuropathy associated with generalized autonomic failure. Small unmyelinated nerves are involved early and prominently in early-onset Val30Met ATTRv, whereas other ATTRv and light-chain amyloidosis often present with large- and small-fiber involvement. Nerve conduction studies, quantitative sudomotor axon testing, and intraepidermal nerve fiber density are useful tools to evaluate denervation. Amyloid deposition can be demonstrated by tissue biopsy of the affected organ or surrogate site, as well as bone-avid radiotracer cardiac imaging. Treatment of light-chain amyloidosis has been revolutionized by monoclonal antibodies and stem cell transplantation with improved 5-year survival up to 77%. Novel gene therapy and transthyretin stabilizers have revolutionized treatment of ATTRv, improving the course of neuropathy (less change in the modified Neuropathy Impairment Score + 7 from baseline) and quality of life. With great progress in amyloidosis therapies, early diagnosis and presymptomatic testing for ATTRv family members has become paramount. ANN NEUROL 2024;96:423-440.

A unique Ca2+-inhibited C-type lectin in shrimp Fenneropenaeuschinensis

C-type lectins (CTLs) are glycan-binding pattern recognition receptors (PRRs) that can bind to carbohydrates on pathogen surfaces, triggering immune responses in shrimp innate immunity. In this study, a unique Ca2+-inhibited CTL named FcLec was identified and characterized in Chinese shrimp Fenneropenaeus chinensis. The full-length cDNA sequence of FcLec was 976 bp (GenBank accession number KU361826), with a 615 bp open reading frame (ORF) encoding 204 amino acids. FcLec possesses a C-type lectin-like domain (CTLD) containing four conserved cysteines (Cys105, Cys174, Cys192, and Cys200) and two sugar-binding site structures (QPD and LNP). The tertiary structure of FcLec deduced revealed three α-helices and eight β-pleated sheets. The mRNA expression levels of FcLec in hemocytes and the hepatopancreas were markedly elevated after stimulation with Vibrio anguillarum and white spot syndrome virus (WSSV). The recombinant FcLec protein exhibited Ca2+-independent hemagglutination and bacterial agglutination, but these activities were observed only in the presence of EDTA to chelate metal ions. These findings suggest that FcLec plays important and functionally distinct roles in the shrimp's innate immune response to bacteria and viruses, enriching the current understanding of the relationship between CTL activity and Ca2+ in invertebrates.

Macromolecules with predominant β-pleated sheet proteins in extracellular vesicles released from Raphanus sativus L. var. caudatus Alef microgreens induce DNA damage-mediated apoptosis in HCT116 colon cancer cells

Plant-derived bioactive macromolecules (i.e., proteins, lipids, and nucleic acids) were prepared as extracellular vesicles (EVs). Plant-derived EVs are gaining pharmaceutical research interest because of their bioactive components and delivery properties. The spherical nanosized EVs derived from Raphanus sativus L. var. caudatus Alef microgreens previously showed antiproliferative activity in HCT116 colon cancer cells from macromolecular compositions (predominantly proteins). To understand the mechanism of action, the biological activity studies, i.e., antiproliferation, cellular biochemical changes, DNA conformational changes, DNA damage, apoptotic nuclear morphological changes, apoptosis induction, and apoptotic pathways, were determined by neutral red uptake assay, synchrotron radiation-based Fourier transform infrared microspectroscopy, circular dichroism spectroscopy, comet assay, 4′,6-diamidino-2-phenylindole (DAPI) staining, flow cytometry, and caspase activity assay, respectively. EVs inhibited HCT116 cell growth in concentration- and time-dependent manners, with a half-maximal inhibitory concentration of 675.4 ± 33.8 μg/ml at 48 h and a selectivity index of 1.5 ± 0.076. HCT116 treated with EVs mainly changed the cellular biochemical compositions in the nucleic acids and carbohydrates region. The DNA damage caused no changes in DNA conformation. The apoptotic nuclear morphological changes were associated with the increased apoptotic cell population. The apoptotic cell death was induced by both extrinsic and intrinsic pathways. EVs have potential as antiproliferative bioparticles.

Clinical Characterization of Hepatic Light Chain Amyloidosis in the Chinese Population: A Single-Center Retrospective Analysis

CONCLUSION Hepatic light chain amyloidosis is caused by malignant plasma cells that abnormally secrete immunoglobulin light chains, which undergo a conformational change to a β-pleated sheet configuration and are deposited in the liver. The amyloid fibrils are deposited mainly in the hepatic sinusoids, the space of Disse and the walls of blood vessels. Previous autopsy studies have shown that the proportion of systemic light chain amyloidosis with liver involvement is extremely high; however, there are fewer systematic analyses of the clinical features in the Chinese population. Therefore, this study retrospectively analysed the clinical data of patients with hepatic lightchain amyloidosis in our center to further explore the clinical characteristics of this disease. A total of 40 patients with hepatic light chain amyloidosis were included in the analysis from 2007 to 2022. Inclusion criteria were positive liver tissue biopsy or positive biopsy of other organs with alkaline phosphatase (AKP) greater than 1.5 times the upper limit of normal or total liver span >15 cm in the absence of heart failure. The median age of these patients was 58±10 years and 80% were male. Of these 40 patients, 66% had light-chain amyloidosis secondary to myeloma and the remainder had primary amyloidosis. The proportion of patients with symptoms related to liver injury as the initial clinical presentation was 33%. Symptoms included malaise, abdominal distention, or jaundice. 40% of patients with symptoms related to abnormal heart function, such as chest tightness, severe shortness of breath, and dizziness. 25% of the patients were first seen for symptoms related to impaired renal function, such as proteinuria with lower limb edema or hypourocrinia. κ-type was found in 28% of the patients and λ-type in 73%. All patients had other sites of involvement, 95% with cardiac involvement (72.9%, Mayo 2004 stage III) and 48% with renal involvement. AKP was elevated in 100% of patients, r-GT in 95%, liver enzymes in 33%, total bilirubin in 44%, and the maximum anteroposterior diameter of the right lobe of the liver was 16.6±1.7 cm (upper limit of normal was 10 cm). In this cohort of patients, 80% received a 2- to 3-drug combination regimen based on proteasome inhibitors and 5% received a 2-drug combination regimen based on melphalan, with a median survival time of 6.5±31.3 months, and patients with a normal total bilirubin level had an overall survival time 2.4 times longer than those with an elevated level (p< 0.05,Figure 1). Contrary to previous reports of hepatic light chain amyloidosis, λ light chain amyloidosis was predominant in our center; elevated total bilirubin was an independent risk factor for survival.

Crotonaldehyde induced structural alterations in Low-Density Lipoprotein: Immunogenicity of the modified protein in experimental animals and auto-antibodies generation in various cancers

Crotonaldehyde (CA), a prominent component of cigarette smoke (CS) is a pervasive environmental pollutant that is a highly toxic, unsaturated aldehyde. Exposure to CA-rich pollutants has been linked to the emergence of many malignancies in humans. To better understand the role of CA in biomolecule modification, this study investigated the detailed structural alterations in low-density lipoprotein (LDL) modified by CA, as well as the immunogenicity of the modified protein in experimental animals and the search for autoantibodies in various cancers patients.In vitro, results indicated alterations in secondary and tertiary structures; examined using UV–visible, fluorescence, far-UV circular dichroism, and Fourier transform infrared spectroscopy techniques. Changes in the oxidation status of LDL were studied by carbonyl content assay and NBT assay. ThT binding assay, scanning, and transmission electron microscopy were used to study aggregate formation. The findings revealed significant structural damage in LDL modified by CA. The modification resulted in the unmasking of hydrophobic clusters, the loss of the protein α-helix, and the formation of β-pleated sheet structure. The amyloid aggregate formation was confirmed through ThT microscopy and electron spectroscopy. Rabbits immunized with crotonaldehyde; lead to structural changes in the LDL; that acted as extra antigenic determinants, eliciting strong antibody response. Immunoglobulin response is highly specific for modified LDL as demonstrated by the ELISA. The presence of antibodies against CA-modified LDL was confirmed by the immunoglobulin content of blood sera from human subjects with lung cancer, and competitive ELISA demonstrated the specificity of these antibodies. This study offers insights into the CA-mediated LDL modification and immunogenicity in lung cancer that will have diagnostic importance.

Topical dexpanthenol effects on physiological parameters of the stratum corneum by Confocal Raman Microspectroscopy.

Topical use of dexpanthenol presents well-established moisturizing properties and maintenance and repair of the skin barrier function, however, its exact action mechanisms are not completely elucidated. In this context, Confocal Raman Microspectroscopy is an optical method that enables non-invasive and non-destructive in vivo analysis with the sensitive acquisition of molecular changes in different skin layers. Herein, the aim was to evaluate the effects of topical dexpanthenol on the components and physiological parameters of the stratum corneum (SC). Ten healthy female subjects underwent skin evaluation by means of a Confocal Raman Spectrometer Skin Analyzer 3510. Spectral data were obtained from the skin of the anterior forearm region, before and 2h after applying a cosmetic formulation containing or not containing 5% dexpanthenol. Semiquantitative analysis of the natural moisturizing factor showed a significant decrease in content after 2 h of topical dexpanthenol application, while the analysis of the lamellar organization of intercellular lipids and the secondary structure of keratin showed a significant increase in hexagonal organization of lipids at the first half of the SC and a significant increase in β-pleated sheet conformation of keratin. Effects of topical dexpanthenol on SC suggest a contribution in increasing fluidity of both lipidic and protein components of the SC and are compatible with dexpanthenol activity in maintaining adequate physiological conditions and preventing transepidermal water loss. This study also contributes to the elucidation of action mechanisms and other concurrent biochemical processes.

COVID-19 Infection and Vaccination and Its Relation to Amyloidosis: What Do We Know Currently?

Amyloidosis is a complex disorder characterized by deposited insoluble fibrillar proteins which misfold into β-pleated sheets. The pathogenesis of amyloidosis can vary but can be the result of immune dysregulation that occurs from sustained high inflammatory states, often known as AA amyloidosis. Multi-organ involvement including hepatic, gastrointestinal, renal, cardiac and immunological pathological manifestations has been observed amongst individuals presenting with amyloidosis. The recent global pandemic of severe acute respiratory syndrome coronavirus 2, also referred to as coronavirus 2019 (COVID-19), has been shown to be associated with multiple health complications, many of which are similar to those seen in amyloidosis. Though COVID-19 is recognized primarily as a respiratory disease, it has since been found to have a range of extra-pulmonary manifestations, many of which are observed in patients with amyloidosis. These include features of oxidative stress, chronic inflammation and thrombotic risks. It is well known that viral illnesses have been associated with the triggering of autoimmune conditions of which amyloidosis is no different. Over the recent months, reports of new-onset and relapsed disease following COVID-19 infection and vaccination have been published. Despite this, the exact pathophysiological associations of COVID-19 and amyloidosis remain unclear. We present a scoping review based on our systematic search of available evidence relating to amyloidosis, COVID-19 infection and COVID-19 vaccination, evaluating current perspectives and providing insight into knowledge gaps that still needs to be addressed going forward.

Structural Modelling of Platelet Activating Factor Acetyl Hydrolase in Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei: Implications on Therapeutics for Leishmaniasis, Chagas Disease, and Sleeping Sickness.

Leishmaniasis, Chagas disease, and sleeping sickness are caused by protozoa Leishmania donovani, Trypanosoma cruzi, and Trypanosoma brucei, respectively. Platelet activating factor acetyl hydrolase (PAF-AH) is an inflammatory protein implicated in pathogenesis of these three infections, thereby making them attractive drug targets. PAF-AH sequences were retrieved from UniProt and aligned using Clustal Omega. Homologous models of parasitic proteins were built based on crystal structure of human PAF-AH and validated using PROCHECK server. Volumes of substrate-binding channel were calculated using the ProteinsPlus program. High throughput virtual screening using Glide program in Schrodinger was done with ZINC drug library against parasitic PAF-AH enzymes. Complexes with best hits were energy-minimized and subjected to 100 ns molecular dynamic simulation and analyzed. PAF-AH enzyme sequences from protozoa Leishmania donovani, Trypanosoma cruzi, Trypanosoma brucei, and human have a minimum of 34% sequence similarity with each other. Corresponding structures show a globular conformation consisting of twisted β-pleated sheets, flanked by α-helices on either side. Catalytic triad of serine-histidine-aspartate is conserved. Substrate-binding channel residues are conserved to an extent, with a lower channel volume in human as compared to target enzymes. Drug screening resulted in identification of three molecules that had better affinities than the substrate to the target enzymes. These molecules fulfill Lipinski's rules for drug likeness and also bind with less affinity to the human counterpart, thereby establishing a high selective index. Structures of PAF-AH from protozoan parasites and humans belong to the same family of enzymes and have a similar three-dimensional fold. However, they show subtle variations in residue composition, secondary structure composition, substrate-binding channel volume, and conformational stability. These differences result in certain specific molecules being potent inhibitors of the target enzymes while simultaneously having weaker binding to human homologue.

Pea protein-xanthan gum interaction driving the development of 3D printed dysphagia diet

With a rapid ageing population, there is an increasing need for the dysphagia diet because of the chewing/swallowing difficulty suffered by the elderly. 3D printing is able to create appetitive and attractive dysphagia diet. We investigated the potential to develop appealing pea protein isolate (PPI) based dysphagia diet using 3D printing by incorporating xanthan gum (XG) at rates of 0.05, 0.1, 0.3, 0.5, 0.7 and 1% (w/w). Their interaction was investigated by scanning electron microscope (SEM), confocal laser scanning microscope (CLSM), Fourier transform infrared (FTIR), and low-field nuclear magnetic resonance (LF-NMR). Rheological properties and 3D printing behavior were also correlated. The 3D printed product was evaluated through International Dysphagia Diet Standardization Initiative (IDDSI) and chewing/swallowing sensory test. Results indicated that, at a relatively small amount of XG addition (0.05, 0.1 and 0.3%), a relatively uniform microstructure was formed with improved mechanical strength (Yield stress, elastic modulus (G′), and viscosity). Further increase in XG addition (0.7 and 1.0%) resulted in an increase in thermodynamic incompatibility between PPI and XG, causing a phase separation and polymer aggregation, which ultimately decreased the mechanical strength of inks. FTIR indicated that the addition of XG increased PPI β-pleated sheet and β-turn, and decreased the contents of random coil and antiparallel β-sheet. XG-0.3% samples demonstrated high printing precision with great self-supporting capability, smooth surface texture and great sensory evaluation, which could be classified as level 4-pureed/extremely thick dysphagia diet. This work provides insights for the development of visually appealing dysphagia diet using 3D printing.

Characterization of freeze-dried oxidized human red blood cells for pre-transfusion testing by synchrotron FTIR microspectroscopy live-cell analysis.

Oxidative treatment of human red blood cells (RBCs) prior to freeze-drying appears to stabilize the RBCs to withstand dried storage at room temperature. To better understand the effects of oxidation and freeze-drying/rehydration on RBC lipids and proteins, single-cell measurements were performed by synchrotron-based Fourier transform infrared (FTIR) microspectroscopy 'live-cell' (unfixed) analysis. Lipid and protein spectral data of tert-butyl hydroperoxide (TBHP)-oxidized RBCs (oxRBCs), FDoxRBCs and control (untreated) RBCs were compared using principal component analysis (PCA) and band integration ratios. The oxRBCs and FDoxRBCs samples had similar spectral profiles that were clearly different to control RBCs. Spectral changes in the CH stretching region of oxRBCs and FDoxRBCs indicated the presence of increased saturated and shorter-chain lipids, consistent with lipid peroxidation and stiffening of the RBC membrane compared to control RBCs. The PCA loadings plot for the fingerprint region of control RBCs corresponding to the α-helical structure of hemoglobin, shows that oxRBCs and FDoxRBCs have conformational changes in the protein secondary structure to β-pleated sheets and β-turns. Finally, the freeze-drying process did not appear to compound or induce additional changes. In this context, FDoxRBCs could become a stable source of reagent RBCs for pre-transfusion blood serology testing. The synchrotron FTIR microspectroscopic live-cell protocol provides a powerful analytical tool to characterize and contrast the effects of different treatments on RBC chemical composition at the single cell level.

Characterization of simplified nonapeptides with broad-spectrum antimicrobial activities as potential food preservatives, and their antibacterial mechanism.

Antimicrobial peptides (AMPs) have attracted attention in the field of food preservatives due to their favorable biosafety and potential antimicrobial activity. However, high synthetic cost, systemic toxicity, a narrow antimicrobial spectrum, and poor antimicrobial activity become the main bottlenecks for their practical applications. To address these questions, a set of derived nonapeptides were designed based on a previously discovered ultra-short peptide sequence template (RXRXRXRXL-NH2) and screened to identify an optimal peptide-based food preservative with excellent antimicrobial properties. Among these nonapeptides, the designed peptides 3IW (RIRIRIRWL-NH2) and W2IW (RWRIRIRWL-NH2) presented a membrane-disruptive and reactive oxygen species (ROS) accumulation mechanism to execute potent and rapid broad-spectrum antimicrobial activity without observed cytotoxicity. Moreover, they exhibited favorable antimicrobial stability regardless of high ionic strength, heat, and excessive acid-base conditions, retaining potent antimicrobial effects for chicken meat preservation. Collectively, their ultra-short sequence length and potent broad-spectrum antimicrobial capacity may be beneficial for the further development of green and safe peptide-based food preservatives.

129. LiaX, a Member of the LiaFSR System, is Essential for Cell Envelope Adaptation System in Enterococcus faecium

Abstract Background Daptomycin (DAP) is an important antibiotic for enterococci. Our previous studies identified LiaX as a surface-exposed protein that is a mediator of cell envelope homeostasis in Enterococcus faecalis (Efs) upon exposure to DAP via activation of the LiaFSR system. LiaX encodes for a soluble protein with an N-terminal of α-helices, and a C-terminus which is β-pleated sheets. Its role in E. faecium, a species of clinical relevance, remains unknown. In this work, we aim to elucidate the function of LiaX of E. faecium (Efm). Methods We used the commensal strain of Efm TX1330RF (DAP minimum inhibitory concentration [MIC] 3 μg/ml), its ΔliaR derivative (TX1330RFΔliaR [DAP MIC 0.125 μg/ml]) and targeted the liaX gene for mutagenesis. Using the PheS* counterselection system, we aimed to create a truncation or in-frame deletion of liaX. Mutants were characterized by determination of DAP MIC and visualization of anionic phospholipid domains by 10-N-nonyl-acridine orange (NAO). The nisin-controlled expression vector, pMSP3535, was used to express LiaX from Efm (LiaXTX1330RF) in Efs host OG1RFΔliaX, which lacks liaX. Expression of LiaX of Efs (LiaXOG1RF) was used as a control. Results In the presence of liaR, we were unable to create truncation or in-frame deletion of liaX after many attempts. In Efm TX1330RFΔliaR, we successfully created an in-frame deletion of liaX, TX1330RFΔliaRliaX (DAP MIC 0.125 μg/ml). NAO staining of all strains of TX1330RF, TX1330RFΔliaR and its ΔliaX mutant showed localization of anionic phospholipid domains at septa of the cells. Interestingly, multiple attempts to complement liaR in TX1330RFΔliaRliaX were also futile. NAO staining of Efs OG1RFΔliaX demonstrated that anionic phospholipid microdomains redistributed away from septa. Expression of LiaXOG1RF in Efs OG1RFΔliaX successfully restored microdomains to septal and polar regions of Efs while expression of LiaXTX1330RF did not show any significant difference in the fluorescence staining compared to its parental host (Efs OG1RFΔliaX). Conclusion Our findings suggest that LiaX has divergent functions as a mediator of cell envelope homeostasis in enterococci and may be essential in the Efm redponse to DAP. Efforts to elucidate its role in DAP resistance in Efm are ongoing. Disclosures Cesar A. Arias, MD, PhD, Entasis Phramceuticals: Grant/Research Support|MeMed Diagnostics: Grant/Research Support|Merck: Grant/Research Support.

Head-to-Head Comparison of Tau-PET Radioligands for Imaging TDP-43 in Post-Mortem ALS Brain.

In vivo detection of transactivation response element DNA binding protein-43kDa (TDP-43) aggregates through positron emission tomography (PET) would impact the ability to successfully develop therapeutic interventions for a variety of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). The purpose of the present study is to evaluate the ability of six tau PET radioligands to bind to TDP-43 aggregates in post-mortem brain tissues from ALS patients. Herein, we report the first head-to-head evaluation of six tritium labeled isotopologs of tau-targeting PET radioligands, [3H]MK-6240 (a.k.a. florquinitau), [3H]Genentech Tau Probe-1 (GTP-1), [3H]JNJ-64326067(JNJ-067), [3H]CBD-2115, [3H]flortaucipir, and [3H]APN-1607, and their ability to bind to the β-pleated sheet structures of aggregate TDP-43 in post-mortem ALS brain tissues by autoradiography and immunostaining methods. Post-mortem frontal cortex, motor cortex, and cerebellum tissues were evaluated, and binding intensity was aligned with areas of elevated phosphorylated tau (ptau), pTDP-43, and β-amyloid. Negligible binding was observed with [3H]MK-6240, [3H]JNJ-067, and [3H]GTP-1. While [3H]CBD-2115 displayed marginal specific binding, this binding did not significantly correlate with the distribution of pTDP-43 and AT8 inclusions. Of the remaining ligands, the distribution of [3H]flortaucipir did not significantly correlate to pTDP-43 pathology; however, specific binding trends to a positive relationship with tau. Finally, [3H]APN-1607 relates most strongly to amyloid load and does not indicate pTDP-43 pathology as confirmed by [3H]PiB distribution in sister sections. Our results demonstrate the prominent nature of mixed pathology in ALS, and do not support the application of [3H]MK-6240, [3H]JNJ-067, [3H]GTP-1, [3H]CBD-2115, [3H]flortaucipir, or [3H]APN-1607 for selective imaging TDP-43 in ALS for clinical research with the currently available in vitro data. Identification of potent and selective radiotracers for TDP-43 remains an ongoing challenge.

Extracellular vesicles derived from microgreens of Raphanus sativus L. var. caudatus Alef contain bioactive macromolecules and inhibit HCT116 cells proliferation

Extracellular vesicles (EVs) are phospholipid bilayer vesicles released from cells, containing natural cargos. Microgreens of Raphanus sativus L. var. caudatus Alef were used in this study as the source of EVs. EVs were isolated by differential centrifugation. The physical properties were determined by dynamic light scattering (DLS) and electron microscopy. The biological and chemical composition were studied by Fourier-transform infrared (FTIR) microspectroscopy and high-performance liquid chromatography analysis, respectively. EVs had a median size of 227.17 and 234.90 ± 23.30 nm determined by electron microscopy and DLS, respectively with a polydispersity index of 0.293 ± 0.019. Electron microscopy indicated the intact morphology and confirmed the size. The FTIR spectra revealed that EVs are composed of proteins as the most abundant macromolecules. Using a curve-fitting analysis, β-pleated sheets were the predominant secondary structure. Notably, the micromolecular biomarkers were not detected. EVs exerted anti-cancer activity on HCT116 colon cancer over Vero normal cells with an IC50 of 448.98 µg/ml and a selectivity index of > 2.23. To conclude, EVs could be successfully prepared with a simple and effective isolation method to contain nano-sized macromolecules possessing anti-cancer activity.

Comparative modeling, comparative molecular docking analyses, and revealing of potential binding pockets of MDM-2: A candidate cancer gene

MDM-2 is also known as E3 ubiquitin-protein ligase encoded by Mdm-2. MDM-2 is an important negative regulator of p53 tumor suppressor and performs key function as an inhibitor of p53 transcriptional activation and E3 ubiquitin ligase. MDM-2 also plays significant role in human cancers and therapeutic target. Hundred different structures were predicted through comparative modeling, threading and ab initio approaches followed by the evaluation of predicted structures through various evaluation tools including ERRAT, ProSa-web, Rampage, molprobidity, verify3D and Anolea. The selected 3D structure of MDM-2 showed 13 α- helix chains, 2 β-pleated sheets along with 97.4468% overall quality factor of the predicted structure. Interestingly, it was observed that only 4.5% residues were present in outlier region and the observed errors were fixed. Moreover, 91.1% residues of the selected structure were present in favored region and 8.9% in allowed region having -6.0 Z-score. High throughput virtual screening and comparative molecular docking studies was performed. Four novel compounds have been reported that showed minimum binding energy (-8.1 Kcal/mol) and maximum binding affinity against MDM-2. Molecular docking analyses revealed that Ser154, Arg155, Pro156, Ser157, Lys185, Ser186, Ser188, Ser190, Ile189, Val247, Glu257, Asp173, Glu174, Glu178, Arg161, Ard181, Lys182, Arg183 and His184 residues are significant residues for therapeutic drug targets. The reported compounds showed effective energy scores. In addition, the site-directed mutagenesis may be helpful for further analyses. The reported compounds may act like potent drug compounds against MDM-2.

In Silico Analysis of Plant Flavonoids as Potential Inhibitors of Newcastle Disease Virus V Protein

Newcastle disease is a viral infection causing serious economic losses to the global poultry industry. The V protein of Newcastle disease virus (NDV) is a pathogenicity determinant having various functions such as the suppression of apoptosis and replication of the NDV. This study was designed to assess the resistance potential of plant flavonoids against the V protein of Newcastle disease virus. Sequence analysis was performed using EXPASY and ProtParam tools. To build the three-dimensional structure of V protein, a homology-modeling method was used. Plant flavonoids with formerly reported therapeutic benefits were collected from different databases to build a library for virtual screening. Docking analysis was performed using the modeled structure of V protein on MOE software. Interaction analysis was also performed by MOE to explain the results of docking. Sequence analysis and physicochemical properties showed that V protein is negatively charged, acidic in nature, and relatively unstable. The 3D structure of the V protein showed eight β-pleated sheets, three helices, and ten coiled regions. Based on docking score, ten flavonoids were selected as potential inhibitors of V protein. Furthermore, a common configuration was obtained among these ten flavonoids. The interaction analysis also identified the atoms involved in every interaction of flavonoid and V protein. Molecular dynamics (MD) simulation confirmed the stability of two compounds, quercetin-7-O-[α-L-rhamnopyranosyl(1→6)-β-D-galactopyranoside] and luteolin 7-O-neohesperidoside, at 100 ns with V protein. The identified compounds through molecular docking and MD simulation could have potential as NDV-V protein inhibitor after further validation. This study could be useful for the designing of anti-NDV drugs.

Enzymatic interesterification of palm olein in a continuous packed bed reactor: Effect of process parameters on the properties of fats and immobilized Thermomyces lanuginosus lipase

Palm olein (POL) was enzymatically interesterified in a continuous packed bed reactor (PBR). The effects of interesterification parameters (flow rates of 4.5–40 g/min and temperatures of 50–90 °C) on the properties of interesterified fats and immobilized Thermomyces lanuginosus lipase (Lipozyme ® TL IM) were investigated. Generally, the percentage of saturated fatty acid at sn-2 remained along with higher flow rates (acyl migration inhibited), while the percentage increased along with higher temperatures and lower flow rates (acyl migration promoted). Solid fat content and crystallization rate of the interesterified fats also showed significant modification with the reaction parameters since these properties are directly influenced by the shift in the triacylglycerol compositions. The occurrence of hydrolysis was also evident during the early stage of interesterification attributed to the lipase's water content, which diminished over time as the water concentration decreased. Under prolonged reaction, the lipase's catalytic activity reduced from 236 U to 65 U, and significant changes in the protein's secondary confirmation were observed. After 100 h of reaction, the α-helix structure decreased (from 23.37% to 16.82%) while β-pleated sheet increased (28.10%–39.59%). The increased fluorescence intensity reflected a significant change in the lipase's tertiary structures.

Identification and Characterization of a Double-Stranded RNA Degrading Nuclease Influencing RNAi Efficiency in the Rice Leaf Folder Cnaphalocrocis medinalis.

Rice leaf folder Cnaphalocrocis medinalis is one of the most serious pests of rice in rice-planting regions worldwide. DsRNA-degrading nucleases (dsRNases) are important factors in reducing the efficiency of RNA interference (RNAi) in different insects. In this study, a dsRNase gene from C. medinalis (CmdsRNase) was cloned and characterized. The CmdsRNase cDNA was 1395 bp in length, encoding 464 amino acids. The CmdsRNase zymoprotein contains a signal peptide and an endonuclease NS domain that comprises six active sites, three substrate-binding sites, and one Mg2+-binding site. The mature CmdsRNase forms a homodimer with a total of 16 α-helices and 20 β-pleated sheets. Homology and phylogenetic analyses revealed that CmdsRNase is closely related to dsRNase2 in Ostrinia nubilalis. Expression pattern analysis by droplet digital PCR indicated that the expression levels of CmdsRNase varied throughout the developmental stages of C. medinalis and in different adult tissues, with the highest expression levels in the fourth-instar larvae and the hemolymph. CmdsRNase can degrade dsRNA to reduce the efficiency of RNAi in C. medinalis. Co-silencing of CmCHS (chitin synthase from C. medinalis) and CmdsRNase affected significantly the growth and development of C. medinalis and thus improved RNAi efficacy, which increased by 27.17%. These findings will be helpful for green control of C. medinalis and other lepidopteran pests by RNAi.

Recent advancements toward non-invasive imaging of retinal amyloid-beta for early detection of Alzheimer's disease.

Recent advancements toward non-invasive imaging of retinal amyloid-beta for early detection of Alzheimer's disease.