Search Results (76)

The growing demand for efficient sustainable biocatalysts for industrial applications has driven the exploration of extremozymes from extremophiles, particularly those thriving in geothermal environments. This study aimed to isolate and characterize extracellular amylase-producing thermophilic bacteria from the Nelumwewa geothermal spring in Sri Lanka, an underexplored ecosystem. Among the isolated thermophilic bacterial strains, NW2 isolates exhibited a prominent extracellular amylase activity. Molecular characterization by 16S rRNA gene sequencing confirmed the close phylogenetic relationship between NW2 and Thermaerobacillus caldiproteolyticus, which is well-known for thermostable proteases. Biochemical assays revealed optimal amylase activity of NW2 isolate at 60 °C and pH 8.0, with a crude enzyme activity of 0.85 U/mL. The enzyme demonstrated efficient hydrolysis of raw cassava starch, highlighting its potential for industrial applications in food, biofuel, and detergent industries. This study reports the first T. caldiproteolyticus-like strain from Sri Lanka with significant extracellular amylase activity, emphasizing the biotechnological potential of geothermal springs as sources of novel extremozymes. These findings contribute to the growing repository of thermostable enzymes, highlighting the need for further exploration of Sri Lanka’s geothermal microbial diversity for industrial biocatalysts. Full article

Earthworms are valued as a dietary protein source in many regions. Earthworm protein can yield bioactive peptides, but enzymatic hydrolysis is inefficient by commercial proteases, and bioactivity development is still inadequate. This study developed a novel efficient method for degrading earthworm protein and investigated the lipid-lowering activity and mechanism of earthworm peptides. It was found that combining autolysis and alcalase exhibited a higher hydrolysis degree of earthworm protein of 43.64 ± 0.78% compared to using autolysis or alcalase only. The hydrolysate significantly reduced lipid accumulation in steatotic hepatocytes. LC-MS/MS results showed that the primary lipid-lowering peptides (EWPs) in the hydrolysate were small molecule peptides with molecular weights of 500–1000 Da and chain lengths of 4–7 amino acid residues. Western blot results demonstrated that EWP regulated the expression of lipid metabolism-related proteins, including APOC3, HMGCR, PCSK9, SREBP1, C/EBP-α, NPC1L1, PPAR-γ, and CYP7A1. Transcriptomic analysis and validation experiments indicated that the lipid-lowering activity of EWP was associated with its suppression of inflammatory factors, such as IL-6. This study presents an efficient enzymatic hydrolysis strategy for earthworm protein utilization, laying the foundation for its application in functional foods such as protein supplements, nutraceutical capsules, hypoallergenic infant formulas, and sports nutrition products. Full article

The ongoing threat of viral pandemics such as COVID-19 highlights the urgent need for novel antiviral therapeutics targeting conserved viral proteins. In this study, peptides of 10–30 kDa derived from the marine diatom Phaeodactylum tricornutum were identified as potential inhibitors of SARS-CoV-2 main protease (Mpro), a key enzyme in viral replication. Peptides less than 60 amino acids in length were retrieved from the UniProt database and aligned with reference antiviral sequences using the Biopython pairwise2 algorithm. Six candidates were selected for structural modeling using AlphaFold2 and Swiss-Model, followed by molecular docking using ClusPro2. LigPlot+ was used to assess molecular interactions, while NetMHCpan 4.1 and AVPpred evaluated immunogenicity and antiviral potential, respectively. Molecular dynamics simulations over 100 ns were conducted using OpenMM. These peptides demonstrated stable binding interactions with key catalytic residues of Mpro. Specifically, peptide A0A8J9SA87 interacted with Cys145 and Glu166, while peptide A0A8J9SDW0 exhibited interactions with His41 and Phe140, both of which are known to be essential for Mpro inhibition. Although peptide A0A8J9X3P8 also interacted with catalytic residues, it exhibited greater structural fluctuations during molecular dynamics simulations and achieved lower AVPpred scores, suggesting lower overall antiviral potential. Therefore, A0A8J9SA87 and A0A8J9SDW0 were identified as the most promising candidates. Molecular dynamics simulations further supported the high structural stability of these peptide-Mpro complexes over a 100 ns timescale, reinforcing their potential as effective inhibitors. These findings support P. tricornutum as a valuable source of antiviral peptides and demonstrate the feasibility of in silico pipelines for identifying therapeutic candidates against SARS-CoV-2. Full article

Antimicrobial resistance (AMR) is a global health threat, with methicillin-resistant Staphylococcus aureus (MRSA) being one of the major resistant pathogens. This study reports the isolation of a novel mangrove-derived bacterium, Virgibacillus chiguensis FN33, as identified through genome analysis and the discovery of a new anionic antimicrobial peptide (AMP) exhibiting anti-MRSA activity. The AMP was composed of 23 amino acids, which were elucidated as NH₃-Glu-Gly-Gly-Cys-Gly-Val-Asp-Thr-Trp-Gly-Cys-Leu-Thr-Pro-Cys-His-Cys-Asp-Leu-Phe-Cys-Thr-Thr-COOH. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for MRSA were 8 µg/mL and 16 µg/mL, respectively. FN33 AMP induced cell membrane permeabilization, suggesting a membrane-disrupting mechanism. The AMP remained stable at 30–40 °C but lost activity at higher temperatures and following exposure to proteases, surfactants, and extreme pH. All-atom molecular dynamics simulations showed that the AMP adopts a β-sheet structure upon membrane interaction. These findings suggest that Virgibacillus chiguensis FN33 is a promising source of novel antibacterial agents against MRSA, supporting alternative strategies for drug-resistant infections. Full article

Sunflower press cake (SPC), a by-product of the edible oil industry, represents a promising source of plant-based protein. This study aimed to investigate and optimize protein recovery from SPC using conventional (CE) and advanced extraction techniques, including Ultrasound and Microwave-Assisted Extraction (UMAE), Pressurized Liquid Extraction (PLE) and Enzyme-Assisted Extraction (EAE). The protein content both in extracts and in the precipitated mass was measured through Lowry assay, while the amino acid profile of the extracted proteins under optimal conditions was analyzed via High-Performance Liquid Chromatography (HPLC). Extraction parameters were optimized using response surface methodology (RSM) for each method. Among the novel methods studied, UMAE and PLE demonstrated superior efficiency over CE, yielding higher protein recovery in significantly shorter extraction times. Optimal UMAE conditions (10 min, 0.03 g/mL, 450 W microwave power, and 500 W ultrasound power) yielded a precipitation yield (PY) of 21.2%, protein recovery in extract (PR_E) of 79.9%, and protein recovery in precipitated mass (PR_P) of 66.3%, with a protein content (PC_P) of 902.60 mg albumin eq./g. Similarly, optimal PLE conditions (6 min, 0.03 g/mL, and 50 °C) resulted in PY, PR_E, and PR_P of 17.7, 68.9, and 47.4%, respectively, with a PC_P of 932.45 mg albumin eq./g. EAE using Aspergillus saitoi protease was comparatively less effective. The amino acid profiling confirmed SPC as a valuable protein source, with glutamic acid, arginine, and aspartic acid being the most abundant. These results highlight the potential of UMAE and PLE as efficient strategies for valorizing edible oil industry by-products into high-quality protein ingredients for food and biotechnological applications. Full article

Soda lakes are extreme saline–alkaline environments that harbor metabolically versatile microbial communities with significant biotechnological potential. This study employed shotgun metagenomics (NovaSeq PE150) to investigate the functional diversity and metabolic potential of microbial communities in Ethiopia’s Chitu and Shala Lakes. An analysis of gene content revealed 554,609 and 525,097 unique genes in Chitu and Shala, respectively, in addition to a substantial fraction (1,253,334 genes) shared between the two, underscoring significant functional overlap. Taxonomic analysis revealed a diverse phylogenetic composition, with bacteria (89% in Chitu Lake, 92% in Shala Lake) and archaea (4% in Chitu Lake, 0.8% in Shala Lake) as the dominant domains, alongside eukaryotes and viruses. Predominant bacterial phyla included Pseudomonadota, Actinomycetota, and Gemmatimonadota, while Euryarchaeota and Nitrososphaerota were prominent among archaea. Key genera identified in both lakes were Nitriliruptor, Halomonas, Wenzhouxiangella, Thioalkalivibrio, Aliidiomarina, Aquisalimonas, and Alkalicoccus. Functional annotation using the KEGG, eggNOG, and CAZy databases revealed that the identified unigenes were associated with various functions. Notably, genes related to amino acid, carbohydrate, and energy metabolism (KEGG levels 1–2) were predominant, indicating that conserved core metabolic functions are essential for microbial survival in extreme conditions. Higher-level pathways included quorum sensing, two-component signal transduction, and ABC transporters (KEGG level 3), facilitating environmental adaptation, stress response, and nutrient acquisition. The eggNOG annotation revealed that 13% of identified genes remain uncharacterized, representing a vast untapped reservoir of novel enzymes and biochemical pathways with potential applications in biofuels, bioremediation, and synthetic biology. This study identified 375 unique metabolic pathways, including those involved in pyruvate metabolism, xenobiotic degradation, lipid metabolism, and oxidative stress resistance, underscoring the microbial communities’ ability to thrive under fluctuating salinity and alkalinity. The presence of carbohydrate-active enzymes (CAZymes), such as glycoside hydrolases, polysaccharide lyases, and oxidoreductases, highlights their role in biomass degradation and carbon cycling. Enzymes such as alkaline proteases (Apr), lipases (Lip), and cellulases further support the lakes’ potential as sources of extremophilic biocatalysts. These findings position soda lakes as reservoirs of microbial innovation for extremophile biotechnology. Future research on unannotated genes and enzyme optimization promises sustainable solutions in bioenergy, agriculture, and environmental management. Full article

Adipocytes secrete adipokines, bioactive molecules crucial for various physiological processes, such as enhancing insulin sensitivity, promoting wound healing, supporting hair growth, and exhibiting anti-aging effects on the skin. With the growing global demand for sustainable and alternative protein sources, insect-derived proteins, particularly from Tenebrio molitor (mealworms), have gained attention due to their high nutritional value and functional bioactivities. This study aims to explore the potential of mealworm-derived protein hydrolysates as novel bioactive materials for promoting adipogenesis and improving adipokine expression, with applications in metabolic health and skin regeneration. Protein hydrolysates (<1 kDa) were prepared using enzymatic hydrolysis with three proteases (alcalase, flavourzyme, and neutrase) and evaluated for their adipogenic activity in 3T3-L1 preadipocytes. Among them, the flavourzyme-derived hydrolysate (Fh-T) exhibited the most significant effects, enhancing adipogenic differentiation and lipid accumulation. Fh-T facilitated adipogenesis by promoting mitotic clonal expansion (MCE) during the early stage of differentiation, which was associated with the upregulation of C/EBPδ and the downregulation of p27. These findings underscore the potential of mealworm-derived protein hydrolysates, particularly Fh-T, as sustainable and functional ingredients for use in glycemic control, skin health, and tissue regeneration. This study provides valuable insights into the innovative use of alternative protein sources in functional foods and cosmeceuticals. Full article

Background/Objectives: Astragalus L. is a genus of the Fabaceae family, encompassing over 3000 species globally, with 380 species found in Turkey. This is the inaugural examination of the phytochemical, antioxidant, antibacterial, and cytotoxic properties of Astragalus pisidicus. Methods: The water and methanolic fractions of four parts (stems, flowers, leaves, root) as well as the whole plant were quantified and identified by Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry (LC–ESI–MS/MS) analysis. Cell death was assessed using the WST-1 assay, while apoptosis was identified by colorimetric protease assay for caspase 2, -3, -6, -8, and -9, as well as cellular DNA fragmentation assay. Antioxidant activity of A. pisidicus water and methanolic extracts was investigated with eight different assays. Antimicrobial activities of the extracts were evaluated against 16 bacterial strains by disc diffusion and broth microdilution methods. Results: A total of 13 phytochemicals were detected in the extracts at various concentrations. Hesperidin (147–40,174 µg/g extract) and hyperoside (363–2677 µg/g extract) comprised the principal constituents among the extracts. Fm (IC₅₀ = 9.57 µg/mL), Rm (IC₅₀ = 14.89 µg/mL), and Sm (IC₅₀ = 9.57 µg/mL) were evaluated as active crude extracts on H1299, HT-29, and Panc-1 cells, while Rm (IC₅₀ = 32.057 µg/mL) and Fm (IC₅₀ = 64.25 µg/mL) were assessed as moderately active on MCF-7 and 22RV1 cells, respectively. The elevation of caspase 2, 3, 6, 8, and 9 enzyme activities, along with DNA fragmentation, signifies that the mode of cell death is apoptosis. According to the disc diffusion test results, Fm, Lm, Sm, and WPm extracts exhibited antimicrobial activity against gram (+) bacteria. Conclusions: A. pisidicus elicited apoptotic cell death in cancer cells selectively by the activation of caspases and subsequent DNA fragmentation and may serve as a novel source of an apoptosis-inducing anticancer drug. Full article

Marine-derived foods, often called blue foods, are promising sustainable alternatives to conventional food sources owing to their abundant amino acids and high protein content. Current treatments for hyperuricemia, a chronic condition attributed to purine metabolism disorders, are associated with various side effects. Novel peptide xanthine oxidase inhibitors have been discovered in the hydrolyzed products of marine fish and invertebrate proteins, which have demonstrated promising therapeutic potential by reducing uric acid levels in vitro and in vivo. This review explores the potential therapeutic effects of xanthine oxidase inhibitors derived from marine fish and invertebrates, summarizes the methods for extracting bioactive peptides from marine organisms, and emphasizes the impact of different proteases on the structure–activity relationship of bioactive peptides. The hypouricemic effects of these bioactive peptides warrant further verification. There is consensus on the in vitro chemical methods used to verify the xanthine oxidase inhibitory effects of these peptides. Considering several cell and animal model development strategies, this review summarizes several highly recognized modeling methods, proposes strategies to improve the bioavailability of bioactive peptides, and advocates for a diversified evaluation system. Although the screening and evaluation methods for antihyperuricemic peptides have been shown to be feasible across numerous studies, they are not optimal. This review examines the deficiencies in bioavailability, synthesis efficiency, and evaluation mechanisms in terms of their future development and proposes potential solutions to address these issues. This review provides a novel perspective for the exploration and application of marine-derived hypouricemic bioactive peptides. Full article

Crustins are a family of antimicrobial peptides (AMPs) that play a pivotal role in the innate immune system of crustaceans. The discovery of novel AMPs from natural sources is crucial for expanding our current database of these peptides. Here, we identified and characterized a novel member of the crustin family, named PpCrus-SWD1, derived from Pollicipes pollicipes. PpCrus-SWD1 consists of 138 amino acids and contains eight cysteine residues that form a conserved ‘four-disulfide core’ structure. Our recombinant PpCrus-SWD1 (rPpCrus-SWD1) exhibited potent inhibitory activity against three Gram-positive bacteria (Staphylococcus aureus, Bacillus sp. T2, and Streptococcus agalactiae) and six Gram-negative bacteria (Aeromonas hydrophila, Escherichia coli, Vibrio anguillarum, Vibrio alginolyticus, Vibrio parahemolyticus, and Acinetobacter sp. L3), with minimum inhibitory concentrations ranging from 16 to 64 μM. Furthermore, rPpCrus-SWD1 demonstrated binding affinity towards both bacteria and pathogen-associated molecular patterns (PAMPs), and damaged bacterial barrier. Additionally, it effectively inhibited alkaline protease activity in S. aureus and V. alginolyticus strains. These findings highlight the potential utility of this newly discovered crustin as an effective alternative to antibiotics. Full article

Chikungunya and Mayaro fevers are viral infectious diseases characterized by fever and arthralgia, for which there are currently no effective vaccines or treatments. The urgent need for novel antiviral agents against Chikungunya virus (CHIKV) and Mayaro virus (MAYV) has led to interest in plant-based compounds that can disrupt the viral replication cycle. Chiococca alba (L.) Hitchc., a Neotropical plant traditionally used by Yucatec Maya healers as an antipyretic and antirheumatic, may hold potential as a source of antiviral agents. This study aimed to evaluate the antiviral potential of C. alba methanolic extracts (CAH21 and CAH24) against CHIKV and MAYV through preliminary in vitro and in silico analyses. The cytotoxicity of two methanolic extracts from C. alba roots was assessed in Vero cells using the neutral red assay, and their viral activity was determined via plaque assay post-treatment. Given the observed antiviral effects, we used computational predictions to explore interactions between the multifunctional nsP2 proteases and secondary metabolites identified in C. alba extracts. The metabolites were identified using high-performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC-MS). Phytochemical analysis revealed the presence of flavonoids, coumarins, and phenolic acids in the C. alba extracts. In vitro assays demonstrated that both extracts inhibited over 70% of activity against CHIKV and MAYV at a concentration of 60 µg/mL. In silico predictions suggested that the flavonoids naringin and vitexin had the highest affinity for the nsP2 proteases of CHIKV and MAYV, indicating their potential as viral inhibitors. Our findings revealed that C. alba extract represents a promising source of novel antiviral compounds. Full article

The skin of amphibians is a rich source of peptides with a wide range of biological activities. They are stored in secretory granules in an inactive form. Upon stimulation, they are secreted together with proteases into the skin. Once activated, they rapidly exert their biological effects, including fighting microorganisms and predators, while their excess is immediately destroyed by the released proteases. To keep bioactive peptides in their initial form, it is necessary to inhibit these enzymes. Several inhibitors for this purpose have previously been mentioned; however, there has not been any reliable comparison of their efficiency so far. Here, we studied the efficiency of methanol and hydrochloric and formic acids, as well as phenylmethylsulfonyl fluoride, in the inhibition of nine frog peptides with the known sequence, belonging to five families in the secretion of Pelophylax esculentus. The results demonstrated that methanol had the highest inhibitory efficiency, while phenylmethylsulfonyl fluoride was the least efficient, probably due to its instability in aqueous media. Possible cleavages between certain amino acid residues in the sequence were established for each of the inhibitors. These results may be helpful for future studies on the nature of proteases and on prediction of the possible cleavage sites in novel peptides. Full article

Microbial communities have been documented as playing many pivotal roles, and contributing to the growth or health performance of animal hosts. Thus, many studies are currently looking for potential beneficial bacteria “probiotics” from diverse environments, including wild species. The present study aimed to investigate the diversity and potential metabolic functions of bacterial communities in the gastrointestinal tract of wild spiny lobsters. The gastrointestinal (GI) tracts of two wild lobster species (Panulirus ornatus and Panulirus homarus) were dissected aseptically and analyzed through high-throughput sequencing, followed by PICRUSt analysis. The results exposed that the most dominant phyla inhabiting both lobster species at the post-puerulus and juvenile stages were Proteobacteria, Firmicutes, Bacteriodota, Patescibacteria, and Verrucomicrobiota, while at the genus level, the GI tracts were mostly dominated by Photobacterium, Candidatus Bacillopora, Vibrio, and Catenococcus at the post-peurulus stage, and Vibrio, Catenococcus, Acanthopleuribacter, Acinetobacter, Pseudoalteromonas, Grimontia, and Photobacterium at the juvenile stage. Further metagenomic prediction analysis discovers many potential probiont properties indicated by the detection of marker genes corresponding to many important metabolic activities, such as antimicrobial compounds (streptomycin, vancomycin, carbapenem, tetracycline, novobiocin, penicillin, cephalosporin, ansamycines, butirosin, and neomycin), antioxidants (e.g., flavonoids and carotenoids), and several important digestive enzymes (e.g., lipase, protease, and amylase). These results suggest that GI tracts of wild spiny lobsters are potential sources to discover novel probionts for aquaculture purposes. Further studies, such as the isolation of the natural product-producing bacteria, or cloning of the beneficial compound-identified genes, are highly recommended to develop novel probiotic strains for aquaculture purposes. Full article

Global public health is facing a major issue with emerging resistance to antimicrobial agents. Antimicrobial agents that are currently on the market are strong and efficient, but it has not been ruled out that these medications will eventually cause resistance to bacteria. Exploring novel bioactive compounds derived from natural sources is therefore, crucial to meet future demands. The present study evaluated the mode of action of the antimicrobial potential protease enzyme SH21. Protease SH21 exhibited antimicrobial activity, strong heat stability (up to 100 °C), and pH stability (pH 3.0 to 9.0). In terms of mode of action, we found that protease SH21 was able to disrupt the bacterial cell membrane as the results of the nucleotide leakage and cell membrane permeability assay. In addition, we also checked inner membrane permeability by PI uptake assay which suggested that protease SH21 has the ability to enter the bacterial cell membrane. Our results revealed that the antimicrobial protease SH21 might be a promising candidate for treating microbial infections. Full article

Recently, there has been a surge towards searching for primitive treatment strategies to discover novel therapeutic approaches against multi-drug-resistant pathogens. Endophytes are considered unexplored yet perpetual sources of several secondary metabolites with therapeutic significance. This study aims to isolate and identify the endophytic fungi from Annona squamosa L. fruit peels using morphological, microscopical, and transcribed spacer (ITS-rDNA) sequence analysis; extract the fungus’s secondary metabolites by ethyl acetate; investigate the chemical profile using UPLC/MS; and evaluate the potential antibacterial, antibiofilm, and antiviral activities. An endophytic fungus was isolated and identified as Aspergillus flavus L. from the fruit peels. The UPLC/MS revealed seven compounds with various chemical classes. The antimicrobial activity of the fungal ethyl acetate extract (FEA) was investigated against different Gram-positive and Gram-negative standard strains, in addition to resistant clinical isolates using the agar diffusion method. The CPE-inhibition assay was used to identify the potential antiviral activity of the crude fungal extract against low pathogenic human coronavirus (HCoV 229E). Selective Gram-positive antibacterial and antibiofilm activities were evident, demonstrating pronounced efficacy against both methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive Staphylococcus aureus (MSSA). However, the extract exhibited very weak activity against Gram-negative bacterial strains. The ethyl acetate extract of Aspergillus flavus L exhibited an interesting antiviral activity with a half maximal inhibitory concentration (IC₅₀) value of 27.2 µg/mL against HCoV 229E. Furthermore, in silico virtual molecular docking-coupled dynamics simulation highlighted the promising affinity of the identified metabolite, orienting towards three MRSA biotargets and HCoV 229E main protease as compared to reported reference inhibitors/substrates. Finally, ADME analysis was conducted to evaluate the potential oral bioavailability of the identified metabolites. Full article