Secondary Structure Analysis Research Articles

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.

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

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.

Capillary Flow-Based One-Minute Quantification of Amyloid Proteolysis.

Quantifying the formation and decomposition of amyloid is a crucial issue in the development of new drugs and therapies for treating amyloidosis. The current technologies for grasping amyloid formation and decomposition include fluorescence analysis using thioflavin-T, secondary structure analysis using circular dichroism, and image analysis using atomic force microscopy or transmission electron microscopy. These technologies typically require spectroscopic devices or expensive nanoscale imaging equipment and involve lengthy analysis, which limits the rapid screening of amyloid-degrading drugs. In this study, we introduce a technology for rapidly assessing amyloid decomposition using capillary flow-based paper (CFP). Amyloid solutions exhibit gel-like physical properties due to insoluble denatured polymers, resulting in a shorter flow distance on CFP compared to pure water. Experimental conditions were established to consistently control the flow distance based on a hen-egg-white lysozyme amyloid solution. It was confirmed that as amyloid is decomposed by trypsin, the flow distance increases on the CFP. Our method is highly useful for detecting changes in the gel properties of amyloid solutions within a minute, and we anticipate its use in the rapid, large-scale screening of anti-amyloid agents in the future.

Molecular insight into binding affinities and blockade effects of selected flavonoid compounds on the PD-1/PD-L1 pathway.

This study investigated the binding mechanisms of the flavonoids apigenin (Api), kaempferol (Kmp), and quercetin (Que) to the PD-L1 dimer using a combination of molecular modeling and experimental techniques. The binding free energy results demonstrated that the flavonoids could tightly bind to the PD-L1 dimer, with the binding abilities following the trend Que > Kmp > Api. Key residues Ile54, Tyr56, Met115, Ala121, and Tyr123 were identified as important for binding. The flavonoids primarily bind to the C-, F-, and G-sheet domains. The spontaneous formation of the complex systems was mainly driven by hydrophobic forces. Dynamic cross-correlation matrix and secondary structure analyses further indicated that the studied flavonoids could stably interact with the binding sites. ELISA results showed that the flavonoids could effectively block PD-1/PD-L1 interactions, although the inhibitory activity of Api was weaker. Therefore, flavonols might be more effective inhibitors compared to flavones. The findings of this study are expected to contribute to the development of novel flavonoids targeting the PD-1/PD-L1 pathway.

Ultrasound-assisted hydrolysis of food waste using glucoamylase: Statistical optimization and mechanistic analysis with molecular simulations

This paper reports investigations in food waste hydrolysis using ternary approach that combines statistical optimization, ultrasound-assisted enhancement of hydrolysis kinetics, and molecular simulations that provide physical insight into the process. Initial optimization of hydrolysis parameters (Box–Behnken design) resulted in total reducing sugar yield of 263.4 mg/g biomass in 42 h. Sonication of hydrolysis mixture at 35 kHz at 20 % duty cycle yielded 4× reduction in hydrolysis time with 22 % enhancement in TRS yield (320 mg/g biomass). Analysis of GLCM's secondary structure through FTIR spectra deconvolution revealed significant changes induced by sonication. Sonication led to reduction in α-helix, and increase in random coil content. Molecular dynamics simulations unveiled majority of amino acid residues associated with GLCM binding pocket in α-helix and random coil regions. Consequently, sonication widened the binding pockets, facilitating easier transport of substrate and product. This effect translated into faster kinetics of enzymatic food waste hydrolysis.

A comprehensive guide for secondary structure and tertiary structure determination in peptides and proteins by circular dichroism spectrometer.

Secondary structure refers to highly regular local sub-structures formed by the polypeptide backbone through hydrogen bonding. The two main types of secondary structures are α-helices and β-strands (which can form β-sheets). The development of a robust circular dichroism (CD) method for structural analysis of biomolecules requires careful consideration of several key factors. Solvent selection plays a crucial role in maintaining the native or desired conformation of the sample while ensuring transparency in the relevant wavelength regions. Aqueous buffers are often preferred for studying proteins in their native state. Optimizing the sample concentration and path length is essential to achieve an optimal absorbance range and maximize the signal-to-noise ratio. Typical concentrations for far-UV CD measurements range from 0.1 to 1mg/ml, with shorter path lengths (1mm) allowing for higher concentrations and longer path lengths (5mm) suitable for dilute solutions. Instrumental parameters, such as scanning speed, accumulations, and nitrogen flow rate, significantly impact the quality and reliability of the acquired CD spectra. Data processing is a critical step in obtaining accurate and interpretable CD spectra. Baseline correction, smoothing, and conversion to mean residue ellipticity are essential for reliable secondary structure analysis.

Potential Transcriptional Enhancers in Coronaviruses: From Infectious Bronchitis Virus to SARS-CoV-2.

Coronaviruses constitute a global threat to human and animal health. It is essential to investigate the long-distance RNA-RNA interactions that approximate remote regulatory elements in strategies, including genome circularization, discontinuous transcription, and transcriptional enhancers, aimed at the rapid replication of their large genomes, pathogenicity, and immune evasion. Based on the primary sequences and modeled RNA-RNA interactions of two experimentally defined coronaviral enhancers, we detected via an in silico primary and secondary structural analysis potential enhancers in various coronaviruses, from the phylogenetically ancient avian infectious bronchitis virus (IBV) to the recently emerged SARS-CoV-2. These potential enhancers possess a core duplex-forming region that could transition between closed and open states, as molecular switches directed by viral or host factors. The duplex open state would pair with remote sequences in the viral genome and modulate the expression of downstream crucial genes involved in viral replication and host immune evasion. Consistently, variations in the predicted IBV enhancer region or its distant targets coincide with cases of viral attenuation, possibly driven by decreased open reading frame (ORF)3a immune evasion protein expression. If validated experimentally, the annotated enhancer sequences could inform structural prediction tools and antiviral interventions.