Secondary Structure Analysis Research Articles

Structural and functional consequences of non-synonymous SNPs within the LAMA2 protein: a molecular dynamics perspective

Clinical phenotypic presentations associated with LAMA2 deficiency have shown a variety of manifestations. LAMA2 mutations are mainly linked to congenital muscular dystrophy, but there is also mounting evidence suggesting their presence in inflammatory breast cancer, laryngopharyngeal squamous cell carcinoma, and ventricular tachycardia related to coronary artery disease and cardiomyopathy. This study examined the structural and functional impacts of 144 non-synonymous single nucleotide polymorphisms (nsSNPs) within the LAMA2 gene. Through multi-tiered sequence and structure-based methods, 11 deleterious and destabilizing mutations were identified (A1362T, E1308Q, E1360G, I1276S, L1195P, M1359T, P1232H, P1238A, P1272L, Y1234H, Y1338C). Further, four mutations (L1195P, Y1234H, P1238A, A1362T), which aligned with conserved positions, were subjected to 500 ns molecular dynamics (MD) simulations. RMSD calculated from MD trajectories highlighted structural disparities between wild-type and mutant forms, with the latter showing greater flexibility. Radius of gyration analysis indicated reduced compactness, solvent accessibility changes suggested unfolding, and hydrogen bond (HB) analysis demonstrated disrupted integrity. The HB analysis revealed disruptions in structural integrity due to diminished hydrogen bonds in mutants. Secondary structure analysis revealed significant alterations in secondary structural content. Principal Component Analysis unveiled increased dynamic behavior in mutants. Gibbs free energy landscape analysis reflected distinct energy minima regions in mutants, indicating structural destabilization. Overall, this study revealed the functional and structural ramifications of nsSNPs in the LAMA2 gene, providing valuable insights into potential disease-causing mutations and warranting future research on understanding LAMA2 associated diseases and disorders.

Identification and Characterization of a Novel Protein Drug Target WP_145928235.1 in Providencia Through Subtractive Proteomics and Molecular Dynamics Simulations

AbstractProvidencia stuartii, the most prevalent strain in its genus, is a significant cause of a range of potentially fatal diseases involving urinary tract infections, lung diseases, and endocarditis, often exhibiting resistance to multiple antibiotics, necessitating alternative therapeutic approaches. Unlike previous efforts focused on known virulence factors, this study delves into the physicochemical and functional characterization of 72 essential hypothetical proteins (EsHPs) from the P. stuartii BE246 genome. Subcellular localization identified 47 cytoplasmic EsHPs as potential drug targets, with the remainder 25 identified as vaccine targets. Subtractive analysis uncovered 68 virulent proteins, 4 of which were nonhomologous to both host and gut microbiota proteomes. Among these, WP_145928235.1 (147 amino acids), a cytoplasmic EsHP, emerged as a novel drug target due to its homologous broad‐spectrum hit. WP_145928235.1 displayed an instability index (II) of 33.14, an aliphatic index of 92.79, and the GRAVY score was ‐0.065 (hydrophilic). Molecular dynamics simulation‐aided conformational study and secondary structure analysis highlighted the protein's flexibility and propensity to interact with inhibitors. Key residues in the 60–90 region, including Asn62, Gly75, Glu81, and Asp85, were critical for interactions and stability, with low frustration indicating residual conservancy. WP_145928235.1 presents a promising target for therapeutic intervention against multidrug‐resistant P. stuartii infections.

Nano-Fe3O4/FeCO3 modified red soil-based biofilter for simultaneous removal of nitrate, phosphate and heavy metals: Optimization, microbial community and possible mechanism

The pollution of nitrogen, phosphorus and heavy metals in surface water is becoming more and more serious, affecting the safety of water quality. In this study, three biofilters were constructed using iron-modified red soil-based filler carriers (RSC, nano-Fe3O4@RSC, and FeCO3@RSC) combined with strain Zoogloea sp. ZP7 to simultaneously remove nitrate (NO3--N), phosphate (PO43--P), copper (Cu2+), and zinc (Zn2+). The long-term operation results showed that the three groups of biofilters could remove 85.0, 90.0, and 89.8% of NO3--N, respectively. Furthermore, the addition of iron compounds enhanced the removal of PO43--P and the resistance to the stress of Cu2+ and Zn2+ in the biofilter. The analysis illustrated that iron modification improved the redox activity and zeta potential of RSC surface. The secondary structure analysis of the protein showed that the microbial secreted proteins were more compact on the surface of the iron-modified RSC, which facilitated the formation of biofilm on the carrier surface. In addition, the iron-modified RSC-based biofilter also showed excellent NO3--N and PO43--P removal efficiency in the treatment of actual surface water. The microbial community analysis results showed that Zoogloea became the dominant species in the biofilter. On the other hand, the presence of iron-reducing bacteria and the expression iron cycle-related genes may contribute to denitrification under low nutrient conditions.

In silico screening of dyes for enhanced silk coloring using the feeding approach

Reducing redundant dyes in the production of colored silk is essential for environmental sustainability. This study aims to innovate silk dyeing processes by employing molecular docking techniques to screen a library of dyes with specific physical properties. For the first time, we identified Acid Blue 25 (Chemical Identifier, CID 22916) and Metanil Yellow (CID 11490) as cost-effective blue and yellow dyes, respectively, that enable the production of intrinsically colored silk by feeding dye-modified mulberry leaves to silkworms. Both dyes demonstrated high selective binding to silk fibroin and retained excellent color performance after degumming. Secondary structural analysis revealed that Acid Blue 25 enhanced mechanical properties by increasing β-sheet content, whereas Metanil Yellow did not. Furthermore, fluorescence spectroscopy indicated that Acid Blue 25 and Metanil Yellow effectively quenched the intrinsic fluorescence of silk fibroin primarily through static quenching, with Acid Blue 25 exhibiting a higher binding constant and more negative binding free energy. Classic molecular dynamic (cMD) simulations revealed that the extensive interaction network of Acid Blue 25, compared to Metanil Yellow, creates a thermodynamically favorable environment for dye-complex binding and stability. Overall, the combined application of virtual screening and the feeding method supports a highly cost-effective and environmentally friendly silk dyeing strategy, with the potential for scaling up in industrial applications.

The effects of ultrasound-assisted glycation on the allergenicity and functional properties of peanut proteins

This study investigated the effects of ultrasound-assisted glycation on the allergenicity and functional properties of peanut proteins. Results showed that ultrasound-assisted glycation increased the degree of glycation reaction of peanut proteins significantly (P < 0.05). ELISA results indicated that the binding of peanut allergens with serum immunoglobulin G (IgG) and immunoglobulin E (IgE) was significantly decreased (P < 0.05). Furthermore, secondary structure analysis revealed a significant increase in β-sheet content, alongside decreases in α-helix, β-turn, and random coil contents (P < 0.05). In addition, intrinsic fluorescence intensity, surface hydrophobicity, and ultraviolet (UV) spectra intensity were diminished (P < 0.05), indicating notable changes in both secondary and tertiary structures of peanut proteins. Moreover, emulsification property, antioxidant activity and in vitro digestibility of peanut proteins showed the most obvious improvements following ultrasound-assisted glycation, and the solubility was increased while turbidity was decreased significantly (P < 0.05). In conclusion, this study demonstrated that ultrasound-assisted glycation not only effectively reduced the allergenicity of peanut allergens, but also improved the overall functional properties of peanut proteins, and the changes in sensitization and functional properties might be closely related to structural changes. This study will provide a theoretical basis for the development of peanut products.

Enhancing salt-induced gelation of egg yolk granules through pH-ultrasound combined treatment: A physicochemical and microstructural analysis

Salted duck eggs are a popular food product, but their high salt content and uneven salt distribution can reduce acceptability. This study investigated the effects of pH-high-intensity ultrasound combined treatment on the salt-induced gelation properties of egg yolk granules. The results showed that the pH5 + 150 W treatment group exhibited the best physical and gelation properties, with the smallest particle size (1597.33 nm), optimal dispersibility (PDI 0.29), and good stability. The gelation properties of this group also demonstrated excellent springiness (0.30 mm), cohesiveness (0.56), and gumminess (0.05 N). Furthermore, the pH5 + 150 W group had the highest water holding capacity of 97.42 % and relatively high hydrophobicity (173.39 μg). Notably, it also showed high oil exudation (2.83 %) and good sandiness (62.07 %). The pH5 + 150 W led to a significant redshift of the fluorescence peak at 335 nm and an enhancement of the peak intensity at 562 nm compared to the control group. Structural characterization revealed a more ordered protein structure and a uniform gel structure with enhanced electrostatic repulsion between oil droplets. Secondary structure analysis of the proteins showed a significant reduction in α-helix and β-sheet, while β-turn and random coil increased. In summary, the pH5 + 150 W treatment displayed the best gelation properties, providing theoretical guidance for enhancing the processing performance of yolk and expanding their application in the food industry.

Identification of novel brain penetrant GSK-3β inhibitors toward Alzheimer’s disease therapy by virtual screening, molecular docking, dynamic simulation, and MMPBSA analysis

One of the significant therapeutic targets for Alzheimer’s disease (AD) is Glycogen Synthase Kinase-3β (GSK-3β). Inhibition of GSK-3β can prevent hyperphosphorylation of tau, and thus prevent formation and accumulation of neurofibrillary tangles and neuropil threads that block intracellular transport, trigger unfolded protein response, and increase oxidative stress, cumulatively leading to neurodegeneration. In this study, we have performed structure-based virtual screening of two small-molecule libraries from ChemDiv CNS databases using AutoDock Vina to identify hit molecules based on their binding affinities compared to that of an established GSK-3β inhibitor, indirubin-3’-monoxime (IMO). Pharmacoinformatic screening on SwissADME and pkCSM servers enabled identification of lead molecules with favorable pharmacoinformatic properties for drug likeliness, including blood brain barrier (BBB) permeability. Further, molecular dynamic simulations identified six candidate lead molecules that show stable complex formation with GSK-3β in dynamic state under physiological conditions. Principal component analysis of the dynamic state was used to plot Free Energy Landscapes (FELs) of GSK-3β-ligand complexes. STRIDE secondary structure analysis of the lowest energy conformations identified from FEL plots, and binding free energy calculations by Molecular Mechanics Poisson-Boltzmann Surface Area ((ΔGbind )MM-PBSA) of the simulation trajectories led to the identification of two ligands as potential lead molecules having favorable free energy landscape profiles as well as significantly lower (ΔGbind )MM-PBSA in dynamic state compared to that of reference inhibitor IMO. Hence, this study identifies two novel brain penetrant GSK-3β inhibitors that are likely to have therapeutic potential against Alzheimer’s disease.

A Double-Stranded Aptamer for Highly Sensitive Fluorescent Detection of Glutathione S-Transferases.

Aptamer-based biosensors have been widely constructed and applied to detect diverse targets. Glutathione S-transferase (GST), a pivotal phase II metabolic enzyme, plays a critical role in biotransformation in vivo, and aberrant GST expression is associated with various health risks. Herein, aptamers targeting GST were systematically selected from a randomized single-stranded DNA (ssDNA) library of 79 nucleotides (nt) using a biotinylated GST-immobilized streptavidin agarose (SA) bead SELEX technology. Following rigorous screening across eight rounds, four aptamers with strikingly similar secondary structures emerged. Among these, Seq3 exhibited the highest affinity towards GST and was selected for further optimization. A semi-rational post-SELEX truncation strategy was then employed based on base composition analysis, secondary structure analysis and affinity assessment. This strategy enabled the systematic removal of redundant nucleotides in Seq3 without compromising its affinity, ultimately yielding a truncated aptamer, Seq3-3, which retains its specificity with a compact 39nt length. Building upon Seq3-3, a double-stranded fluorescent aptamer probe was ingeniously designed for the in vitro detection of GST. The detection mechanism hinges on the competitive displacement of the complementary chain from the probe, mediated by the target protein, leading to the separation of the antisense oligonucleotide from the double-stranded complex. This process triggers the restoration of the fluorescence signal, enabling sensitive detection, and the probe exhibits excellent response within a linear range of GST activity ranging from 0 to 1500 U/L. The results show that not only an efficient strategy for screening robust and practicable aptamers but also an ultrahighly sensitive detection platform for GST was established.

Towards identification of therapeutics against multi-infections and cancers causing Propionibacterium acnes: Molecular modeling and dynamics simulation investigation

Skin conditions related to Propionibacterium acnes are a noteworthy health concern affecting millions globally. However, recent studies suggest that P. acne is also linked to other serious health conditions such as various kinds of cancer such as prostate cancer, breast cancer, and gastric cancer. The P. acnes sialidase enzyme plays an important role in the pathogenesis of this infection by supporting bacterial invasion and adhesion to the host tissues. In the current study, Medicinal Fungi Secondary Metabolites and Therapeutics (MeFSAT) consisting of 1830 compounds were put through comprehensive computational analysis to identify the strongest inhibitors that target the primary pathogenic enzyme that triggers the dermal infection. Upon analyzing docking studies, it is anticipated that most of these compounds are strongly attached to the specific protein vital for bacterial infection based on their binding score (threshold < −7 kcal/mol). The top 3 compounds were selected as MSID000006, MSID000010, and MSID000012 as strong inhibitors based on their binding score. Furthermore, every compound showed an excellent ADMET profile and adhered to Lipinski’s rule. Next, Density Functional Theory (DFT) analysis was carried out to analyze compounds’ optimized structures and electrostatic interactions with high accuracy. Subsequently, molecular dynamics simulations (MDS) were carried out to evaluate compound stability. The particular compounds showed better stability under a dynamic environment. Furthermore, principal component analysis (PCA) and hydrogen bond analysis was carried out to understand the complexity of vast datasets and the stability of the compounds respectively. Moreover, in salt bridge analysis, Asp-Arg were the most observed interactions, suggesting better electrostatic interaction and good stability. Secondary structure analysis was conducted for better comprehension of protein structure. MMPBSA/GBSA analysis was performed to calculate the free energies of simulation trajectories as MSID000006 favorable binding energy as compared to the MSID000010 and MSID000012. WaterSwap validated the MMPBSA findings by calculating the binding free energies of the compounds upon swapping the water molecule with the ligand. In the end, entropy energy was estimated to figure out the disorder in the process. The entropy results showed that MSID00006 has greater stability as compared to MSID000010 due to lesser disorder in the process. Our findings revealed the compounds MSID000006, MSID000010, and MSID000012 could play favorable roles in the management of P. acnes disease by showing their potential as strong inhibitors.

Construction waste as a filler of denitrification biofilter for nitrate utilization from wastewater: Characteristics, performance, microbial community and soilless culture

Construction waste (CW) is produced in large quantities, resulting in severe land occupation and resource depletion. This study utilized CW as fillers to construct a denitrification biofilter (DNBF-CW) for treating secondary effluent from wastewater plants. Performance and mechanism were analyzed by water quality, biomass and its distribution, physicochemical characteristics, microbial community structure, extracellular polymeric substances and protein secondary structure analysis. Results indicated that DNBF-CW achieve NO3−-N and chemical oxygen demand (COD) removal efficiencies (RE) of 97 % and 70 %. The β-sheet of DNBF-CW increased from 47 % to 58 %, accompanied by decrease in random curls from 22 % to 0. Post-use CW showed potential as soilless cultivation substrates, boosting germination rates by 42 ± 7 %. Mechanism investigations elucidated that ZX3 improved efficiency by modulating microbial community composition, with Pseudomonas reaching 37 %. This study shows the multiple use of construction waste, which presents a novel, efficient, and eco-friendly solution for water treatment.

Comprehensive bioinformatics assessments of the ROP34 of Toxoplasma gondii to approach vaccine candidates

IntroductionRhoptries proteins (ROPs) are crucial throughout different stages of the Toxoplasma gondii (T. gondii) lifecycle, playing key roles in both the invasion of host cells and their subsequent survival. ROP34 is particularly noteworthy as it significantly influences host gene expression and aids in the transition from the tachyzoite to the bradyzoite form.Materials and methodsThis research utilized various bioinformatics tools to assess physico-chemical properties, allergenic and antigenic characteristics, sites for post-translational modifications (PTMs) and protein's secondary and three-dimensional structures of the ROP34 protein. Furthermore, the study identified potential B-cell, MHC-binding, and cytotoxic T-lymphocyte (CTL) epitopes within the ROP34 sequence.ResultsThe ROP34 peptide comprised 553 amino acid residues, with a calculated average molecular weight (MW) of 61.60149 kDa, an aliphatic index of 73.98, and a GRAVY score of − 0.554. The antigenicity of the multi-epitope peptide was estimated to be 0.526563 and 0.6025 by the ANTIGENpro and VaxiJen servers, respectively, suggesting ROP34 as an immunogenic protein with no allergenic potential. Secondary structure analysis revealed a composition of 52.80% random coil, 36.17% alpha helix, and 11.03% extended strand. The Ramachandran plot for the refined model depicted that 97.46% of the residues were situated in the favored region.ConclusionThis in silico research serves as a foundation for designing effective immunization tactics to target toxoplasmosis. The present article lays the groundwork for future studies and offers perspectives for the advancement of an appropriate toxoplasmosis vaccine.Graphical

In Vitro Selection and Characterization of a Light-up DNA Aptamer for Thiazole Orange.

A new DNA aptamer that binds to the target Thiazole Orange-biotin (TO1-biotin) was isolated after nine rounds of in vitro selection. The selection was performed on streptavidin-coated beads with the target bound to the surface and with free dye in solution in higher selection rounds to select for slower off-rate binding. Using next-generation sequencing (NGS), the libraries after the 4th and 9th rounds of selection were sequenced to identify enriched sequences. Several sequence families emerged, showing superior fluorescence enhancement and high affinity for the target compared to the other families obtained by NGS analysis. These sequence families were further studied to understand the binding interactions better. Primary sequence and secondary structure analysis tools were used to identify a hypothetical three-tiered G-quadruplex motif for these families. This indicates that the TO1-biotin DNA aptamer identified here uses a similar ligand-binding topology to the original Mango RNA aptamer.

Genetic polymorphism in untranslated regions of PRKCZ influences mRNA structure, stability and binding sites

BackgroundVariations in untranslated regions (UTR) alter regulatory pathways impacting phenotype, disease onset, and course of disease. Protein kinase C Zeta (PRKCZ), a serine-threonine kinase, is implicated in cardiovascular, neurological and oncological disorders. Due to limited research on PRKCZ, this study aimed to investigate the impact of UTR genetic variants’ on binding sites for transcription factors and miRNA. RNA secondary structure, eQTLs, and variation tolerance analysis were also part of the study.MethodsThe data related to PRKCZ gene variants was downloaded from the Ensembl genome browser, COSMIC and gnomAD. The RegulomeDB database was used to assess the functional impact of 5’ UTR and 3’UTR variants. The analysis of the transcription binding sites (TFBS) was done through the Alibaba tool, and the Kyoto Encyclopaedia of Genes and Genomes (KEGG) was employed to identify pathways associated with PRKCZ. To predict the effect of variants on microRNA binding sites, PolymiRTS was utilized for 3’ UTR variants, and the SNPinfo tool was used for 5’ UTR variants.ResultsThe results obtained indicated that a total of 24 variants present in the 3’ UTR and 25 variants present in the 5’ UTR were most detrimental. TFBS analysis revealed that 5’ UTR variants added YY1, repressor, and Oct1, whereas 3’ UTR variants added AP-2alpha, AhR, Da, GR, and USF binding sites. The study predicted TFs that influenced PRKCZ expression. RNA secondary structure analysis showed that eight 5’ UTR and six 3’ UTR altered the RNA structure by either removal or addition of the stem-loop. The microRNA binding site analysis highlighted that seven 3’ UTR and one 5’ UTR variant altered the conserved site and also created new binding sites. eQTLs analysis showed that one variant was associated with PRKCZ expression in the lung and thyroid. The variation tolerance analysis revealed that PRKCZ was an intolerant gene.ConclusionThis study laid the groundwork for future studies aimed at targeting PRKCZ as a therapeutic target.

Characteristics of organic matter driven by Eichhornia crassipes during co-contamination with per(poly)fluoroalkyl substances (PFASs) and microplastics (MPs)

Co-contamination with MPs and PFASs has been recorded, particularly in surface-water environments. Floating macrophyte microcosms are an important part of the surface water ecosystem, and dissolved organic matter (DOM) driven by floating macrophytes (FMDDOM) is critical for maintaining material circulation. However, knowledge gaps remain regarding the impact of MPs and PFASs co-pollution on FMDDOM. An greenhouse simulation experiment was conducted in this study to investigate the effects of four PFASs, perfluorooctanoic acid (PFOA), perfluoro-octane-sulfonic acid (PFOS), perfluoro-2-methyl-3-oxahexanoic acid (Gen X), and potassium 9-chlorohexadecafluoro-3-oxanonane-1-sulfonate (F-53B), on FMDDOM sourced from Eichhornia crassipes (E. crassipes), a typical floating macrophyte, in the presence and absence of polystyrene (PS) MPs. Four PFASs increased FMDDOM release from E. crassipes, leading to a 32.52–77.49 % increase in dissolved organic carbon (DOC) levels. PS MPs further increased this, with results ranging from −21.28 % to 26.49 %. Based on the parallel factor analysis (PARAFAC), FMDDOM was classified into three types of fluorescent components: tryptophan-like, humic-like, and tyrosine-like compounds. Contaminants of MPs and PFASs modified the relative abundance of these three components. Protein secondary structure analysis showed that fluorocarbon bonds tended to accumulate on the α-helix of proteins in FMDDOM. The relative abundance of fluorescent and chromophorous FMDDOMs varied from 0.648 ± 0.044 to 0.964 ± 0.173, indicating that the photochemical structures of the FMDDOM were modified. FMDDOM exhibits decreased humification and increased aromaticity when contaminated with MPs and PFASs, which may be detrimental to the geochemical cycling of carbon. This study offers a theoretical basis for assessing the combined ecological risks of MPs and PFASs in floating macrophyte ecosystems.

Whole genome sequencing and genome characterization of Aichivirus isolated from Korean adults.

The whole-genome sequence (WGS) analysis of Aichivirus (AiV) identified in Korea was performed in this study. Using Sanger and Nanopore sequencing, the 8228-nucleotide-long genomic sequence of AiV (OQ121963) was determined and confirmed to belong to genotype A. The full-length genome of OQ121963 consisted of a 7296 nt open reading frame (ORF) that encodes a single polyprotein, and 5' UTR (676 nt) and 3' UTR (256 nt) at 5' and 3' ends, respectively. The ORF consisted of leader protein (L), structural protein P1 (VP0, VP1, and VP3), and nonstructural protein P2 (2A, 2B, and 2C) and P3 (3A, 3B, 3C, and 3D). The secondary structure analysis of the 5' UTR identified only stem-loop C (SL-C) and not SL-A and SL-B. The variable region of the AiV genome was analyzed by MegAlign Pro and reconfirmed by SimPlot analysis using 16 AiV whole genomes known to date. Among the entire regions, structural protein region P1 showed the lowest amino acid identity (96.07%) with reference sequence AB040749 (originated in Japan; genotype A), while the highest amino acid identity (98.26%) was confirmed in the 3D region among nonstructural protein region P2 and P3. Moreover, phylogenetic analysis of the WGS of OQ121963 showed the highest homology (96.96%) with JX564249 (originated in Taiwan; genotype A) and lowest homology (90.14%) with DQ028632 (originated in Brazil; genotype B). Therefore, the complete genome characterization of OQ121963 and phylogenetic analysis of the AiV conducted in this study provide useful information allowing to improve diagnostic tools and epidemiological studies of AiVs.

A Novel Heterozygous Intronic FBN1 Variant Contributes to Aberrant RNA Splicing in Marfan Syndrome.

Marfan syndrome (MFS) is a complex genetic systemic connective tissue disorder. It is well known that genetic factors play a critical role in the progression of MFS, with nearly all cases attributed to variants in the FBN1 gene. We investigated a Chinese family with MFS spanning two generations. Whole exome sequencing, in silico analysis, minigene constructs, transfection, RT-PCR, and protein secondary structure analysis were used to analyze the genotype of the proband and his father. The main clinical manifestations of the proband and his father were subluxation of the left lens and high myopia with pectus deformity. Whole exome sequencing identified a novel single nucleotide variant (SNV) in the FBN1 gene at a non-canonical splice site, c.443-3C>G. This variant resulted in two abnormal mRNA transcripts, leading to a frameshift and an in-frame insertion. Further invitro experiments indicated that the c.443-3C>G variant in FBN1 was pathogenic and functionally harmful. This research identified a novel intronic pathogenic FBN1: c.443-3C>G gene variant, which led to two different aberrant splicing effects. Further functional analysis expands the variant spectrum and provides a strong indication and sufficient basis for preimplantation genetic testing for monogenic disease (PGT-M).

Secondary structure propensities of the Ebola delta peptide E40 in solution and model membrane environments

The Ebola delta peptide is an amphipathic, 40-residue peptide encoded by the Ebola virus, referred to as E40. The membrane-permeabilising activity of the E40 delta peptide has been demonstrated in cells and lipid vesicles suggesting the E40 delta peptide likely acts as a viroporin. The lytic activity of the peptide increases in the presence of anionic lipids and a disulphide bond in the C-terminal part of the peptide. Previous in silico work predicts the peptide to show a partially helical structure, but there is no experimental information on the structure of E40. Here, we use circular dichroism spectroscopy to report the secondary structure propensities of the reduced and oxidised forms of the E40 peptide in water, detergent micelles, and lipid vesicles composed of neutral and anionic lipids (POPC and POPG, respectively). Results indicate that the peptide is predominately a random coil in solution, and the disulphide bond has a small but measurable effect on peptide conformation. Secondary structure analysis shows large uncertainties and dependence on the reference data set and, in our system, cannot be used to accurately determine the secondary structure motifs of the peptide in membrane environments. Nevertheless, the spectra can be used to assess the relative changes in secondary structure propensities of the peptide depending on the solvent environment and disulphide bond. In POPC-POPG vesicles, the peptide transitions from a random coil towards a more structured conformation, which is even more pronounced in negatively charged SDS micelles. In vesicles, the effect depends on the peptide-lipid ratio, likely resulting from vesicle surface saturation. Further experiments with zwitterionic POPC vesicles and DPC micelles show that both curvature and negatively charged lipids can induce a change in conformation, with the two effects being cumulative. Electrostatic screening from Na+ ions reduced this effect. The oxidised form of the peptide shows a slightly lower propensity for secondary structure and retains a more random coil conformation even in the presence of PG-PC vesicles.

Secondary structure analysis of proteins within the same topology group

The native conformation of a protein plays a decisive role in ensuring its functionality. It is established that the spatial structure of proteins may exhibit a greater degree of conservation than the corresponding amino acid sequences. This study aims to clarify structural distinctions between homologous and non-homologous proteins with identical topology. The analysis focuses on secondary structures with special emphasis on their fraction, distribution along the polypeptide chain, and chirality. Three different groups of proteins with identical topology were considered according to the CATH database: a homologous group of Globins, a group of Phycocyanins, which is often considered as a potential relative of globins, and a diverse assembly of other globin-like proteins. Some structural patterns in the distribution of secondary structure have been identified within Globins. A similar profile was observed in Phycocyanins, in contrast to the third group. In addition, a distinguishable structural motif, including structures such as 310-helix and irregular structure, has been found in both Globins and Phycocyanins, which can be proposed as an evolutionary imprint.

A carboxymethyl cellulase from the yeast Cryptococcus gattii WM276: Expression, purification and characterisation

Cryptococcus gattii and its medical implications have been extensively studied. There is, however, a significant knowledge gap regarding cryptococcal survival in its environmental niche, namely woody material, which is glaring given that infection is linked to environmental populations. A gene from C. gattii (WM276), the predominant global molecular type (VGI), has been sequenced and annotated as a putative cellulase. It is therefore, of both medical and industrial intertest to delineate the structure and function of this enzyme. A homology model of the enzyme was constructed as a fusion protein to a maltose binding protein (MBP). The CGB_E4160W gene was overexpressed as an MBP fusion enzyme in Escherichia coli T7 cells and purified to homogeneity using amylose affinity chromatography. The structural and functional character of the enzyme was investigated using fluorescence spectroscopy and enzyme activity assays, respectively. The optimal enzyme pH and temperature were found to be 6.0 and 50 °C, respectively, with an optimal salt concentration of 500 mM. Secondary structure analysis using Far-UV CD reveals that the MBP fusion protein is primarily α-helical with some β-sheets. Intrinsic tryptophan fluorescence illustrates that the MBP-cellulase undergoes a conformational change in the presence of its substrate, CMC-Na+. The thermotolerant and halotolerant nature of this particular cellulase, makes it useful for industrial applications, and adds to our understanding of the pathogen's environmental physiology.

Discovery of 8-hydroxy-2-quinoline carbaldehyde derivatives as inhibitors for M1 aminopeptidase of Leishmania donovani

M1 aminopeptidase is a metallopeptidase that plays a vital role in protein catabolism and has been identified as a validated drug target in various parasites; however, our understanding of this enzyme is restricted for leishmanial parasite. The present investigation involved the purification of Leishmania donovani M1 aminopeptidase (LdM1AP) to homogeneity by affinity chromatography. Purified LdM1AP was observed to be enzymatically active and displayed maximal activity in the presence of cobalt ions, whereas secondary structure analysis confirmed the dominance of α-helices. Intrinsic fluorescence and quenching studies of LdM1AP has revealed that tryptophan residues were predominantly concealed within the hydrophobic areas. The synthesized 8-hydroxy-2-quinoline carbaldehyde derivatives were screened, wherein HQ2 and HQ12 were found as potent inhibitors for LdM1AP that compete with the substrate and exhibit pharmacokinetic properties as well as no toxicity for macrophages. Moreover, structural insights of protein and ligand complexes demonstrated that lead compounds mostly interact via hydrophobic contacts into the substrate binding pocket of LdM1AP. Furthermore, lead compounds exhibited a greater affinity for LdM1AP compared to the substrate during in vitro and in silico studies. This report establishes the possibility of quinoline derivatives to target the LdM1AP activity and provide a platform to design the specific antileishmanial drugs.