Soluble Protein Research Articles

Quantitative Proteomic Analysis of Lysine Malonylation in Response to Salicylic Acid in the Roots of Platycodon grandiflorus

Salicylic acid, as a plant hormone, significantly affects the physiological and biochemical indexes of soluble sugar, malondialdehyde content, peroxidase, and superoxide dismutase enzyme activity in Platycodon grandiflorus. Lysine malonylation is a post-translational modification that involves various cellular functions in plants, though it is rarely studied, especially in medicinal plants. In this study, the aim was to perform a comparative quantitative proteomic study of malonylation modification on P. grandiflorus root proteins after salicylic acid treatment using Western blot with specific antibodies, affinity enrichment and LC-MS/MS analysis methods. The analysis identified 1907 malonyl sites for 809 proteins, with 414 proteins and 798 modification sites quantified with high confidence. Post-treatment, 361 proteins were upregulated, and 310 were downregulated. Bioinformatics analysis revealed that malonylation in P. grandiflorus is primarily involved in photosynthesis and carbon metabolism. Physiological and biochemical analysis showed that salicylic acid treatment increased the malondialdehyde levels, soluble protein, superoxide dismutase, and peroxidase activity but did not significantly affect the total saponins content in P. grandiflorus. These findings provide an important basis for exploring the molecular mechanisms of P. grandiflorus following salicylic acid treatment and enhance understanding of the biological function of protein lysine malonylation in plants.

Effects of Supplementation of Different Proteins on the Rheological Properties of Liquid Whole Eggs

This study aimed to evaluate the impact of increasing total protein content on the rheological properties of liquid whole eggs. The study fills the gap between fundamental science and industrial application by exploring the potential for developing high-protein, functional egg products for health-conscious consumers and the food industry. Liquid whole egg samples were enriched with different percentages of egg white and whey proteins. Proteins were added either before or after heat treatment, followed by homogenization, to analyze the effects on rheological behavior. Results indicated that whey protein samples exhibit near-Newtonian behavior due to their high solubility and minimal protein interactions, while egg white protein samples, especially at higher concentrations, induce shear thinning behavior and increased viscosity due to their water-binding capacity and partial heat-induced denaturation. Flow behavior index (n) decreased by 62.7 ± 5.7% and 25.2 ± 1.8% for heat treated with 10% added egg white and whey protein samples, respectively; meanwhile, it decreased by 2.64 ± 0.06% for 10% of added egg white protein and increased by 12.0 ± 1.0% for 10% of whey protein when heat treatment was induced prior to protein additions. The findings from this study offer valuable insights for developing functional food enriched with whey and egg white proteins.

Effects of Cryoprotectants on the Physicochemical Properties of Bonito (Sarda sarda) Fillets Subjected to Multiple Freeze–Thaw Cycles

ABSTRACT In this study, bonito (Sarda sarda) fillets were treated with sucrose-sorbitol, sodium tripolyphosphate, and sodium alginate as cryoprotectants before freezing to prevent changes in fish muscle during multiple freezing cycles and frozen storage. Lower soluble protein contents were determined in cryoprotectant-treated groups than in the control group. Lower polar and non-polar free amino acid contents were determined with the sodium tripolyphosphate group in the last cycle compared to the other groups. Cryoprotectant-added fillets had higher water-holding capacity and lower hardness values than the control. It was determined that the most effective cryoprotectant was sodium tripolyphosphate.

The Influence of the Nutritional and Mineral Composition of Vegetable Protein Concentrates on Their Functional Properties

Vegetable proteins derived from legumes, cereals or pseudocereals have increased in popularity in recent years, becoming very interesting for the food industry. In addition to their nutritional interest, these products have techno-functional properties that allow them to be used in the production of a wide variety of foods. This research has studied the nutritional and mineral composition of 12 samples of rice, pea and soy concentrates. The objective was to investigate the influence of this nutritional composition, mainly mineral components, on the techno-functional properties (water- and oil-binding capacity, swelling, emulsifying, gelling and foaming capacities) of these concentrates. For this purpose, a Pearson correlation matrix and a GH biplot method were applied. The results showed that there is a correlation between mineral content and functional properties. Mg, K and Ca were positively correlated with protein solubility index, oil absorption capacity and swelling capacity. Na and P contents were positively related to water absorption capacity and emulsifying capacity. Gelling capacity was positively correlated with Mg contents and negatively correlated with Cu and Fe contents. The preliminary results reported in this study highlight the necessity to further assess the influence of non-protein components on the techno-functionality of protein concentrates.

Exogenous ATP Functions in Alleviating the Decrease in Quality of Grape (Vitis vinifera L.) Fruits After Harvest

ABSTRACTOur current study investigated the effects of exogenous ATP (adenosine triphosphate) on the post‐harvest quality of grape (Vitis vinifera L.) fruits. The results demonstrated that treatment with exogenous ATP effectively alleviated the decreases in fruit firmness, total soluble solids (TSS), vitamin C content and soluble protein of the harvested grape fruits. Furthermore, exogenous ATP also significantly enhanced the content of the titratable acidity (TA) of grape fruits after harvest. This was reflected by the observations that ATP treatment increased the hardness, TSS content, vitamin C content and soluble protein content of the harvested grape fruits by 14.39%, 6.29%, 31.05% and 15.18% on average during 8 days of storage, respectively, compared to the controls. In addition, the harvested grape fruits treated with exogenous ATP had higher activities of SOD (superoxide dismutase) and CAT (catalase) and higher contents of total phenolics and flavonoids compared to the controls. This was reflected by the observations that ATP treatment increased the SOD activity, CAT activity, flavonoid content and total phenol content by 39.79%, 19.43%, 17.74% and 15.06% on average during 8 days of storage, respectively, compared to the controls. These observations highlighted that exogenous ATP can function in alleviating the decrease in quality of grape fruits after harvest and increasing the antioxidative ability of harvested grape fruits during the storage period. Consequently, treatment with exogenous ATP could offer an efficacious approach for preserving the quality of grape fruits during the storage period.

A structural genomics approach to investigate Dystrophin mutations and their impact on the molecular pathways of Duchenne muscular dystrophy

BackgroundDystrophin is a key protein encoded by the DMD gene, serves as a scaffold linking the cytoskeleton to the extracellular matrix that plays a critical role in muscle contraction, relaxation, and structural integrity. Mutations, particularly single-point amino acid substitutions, can lead to dysfunctional Dystrophin, causing muscular dystrophies, with Duchenne muscular dystrophy (DMD) being the most severe form.ObjectiveThis study aimed to evaluate the effects of 184 single-point amino acid substitutions on the structure and function of Dystrophin using computational approaches.MethodsMany computational tools were used to predict the impact of amino acid substitutions on protein stability, solubility, and function. Pathogenic potential was assessed using disease phenotype predictors and CADD scores, while allele frequency data from gnomAD contextualized mutation prevalence. Additionally, aggregation propensity, frustration analysis, and post-translational modification sites were analyzed for functional disruptions.ResultsOf the 184 substitutions analyzed, 50 were identified as deleterious, with 41 predicted to be pathogenic. Seventeen mutations were localized in the Calponin-homology (CH) 1 domain, a critical functional region of Dystrophin. Six substitutions (N26H, N26K, G47W, D98G, G109A, and G109R) were predicted to decrease protein solubility and were located in minimally frustrated regions, potentially compromising Dystrophin functionality and contributing to DMD pathogenesis.ConclusionThis study provides novel insights into the molecular mechanisms of DMD, highlighting specific mutations that disrupt Dystrophin’s solubility and function. These findings could inform future therapeutic strategies targeting Dystrophin mutations to address DMD pathogenesis.

Foliar N Supplementation Improves Rapeseed Transplanting Survival Rate and Yield

Transplanting shock induced by mechanical transplanting technique in dry land has a negative effect on the growth and grain yield of rapeseed (Brassica napus L.). We hypothesized that foliar nitrogen (N) supplementation plays a positive role in improving rapeseed transplanting survival rate and yield. The aim of the study was to elucidatethe morphological and physiological mechanisms of foliar N supplementation in rapeseed responding to transplanting shock. Through a 1-year pot experiment and a 2-year field experiment, foliar N supplementation at concentrations of 0, 2.5, 5.0, and 7.5 g N m−2 was set up to investigate the regulation of foliar N supplementation on the carbon (C) and N structure of seedlings before transplanting. Then we investigated the responses of morphological and physiological changes of shoot, root regeneration ability, transplanting survival rate, and grain yield to transplanting shock. The results showed that foliar N supplementation prior to transplanting increased the short-term activities of GS, GOGAT, and GDH, may improve the conversion of soluble sugar to soluble protein, increased N accumulation, and lowered the C:N ratio of rapeseed seedlings. Rapeseed seedlings with a low C:N ratio showed greater potential for root development after transplanting. Moreover, appropriate foliar N supplementation (2.5–5.0 g N m−2) not only up-regulated the positive feedback of osmoregulatory substances and antioxidant system for improving stress resistance, but also enhanced the synergistic growth of shoot and root by increasing root growth potential during transplanting shock. Therefore, the transplanting survival rate increased by 13.81–19.20%, and the grain yield increased by 25.15–30.56%. The optimal foliar N supplementation before transplanting may be used as a simple and effective agricultural measure to alleviate the negative effects of transplanting shock on rapeseed.

Stress-Induced Response and Adaptation Mechanisms in Bacillus licheniformis PSKA1 Exposed With Abiotic and Antibiotic Stresses.

Soil ecosystems consist of diverse microbial communities with great potential for ecological and biotechnological applications. These communities encounter various abiotic stresses, which expedite the activation of transient overexpression of heat shock proteins (HSPs). In the present study, a soil bacterium was isolated and identified as Bacillus licheniformis strain PSK.A1, and its growth parameters were optimized before exposing it to heat, salt, pH, and antibiotic stress conditions. Comparative protein expression was analyzed using SDS-PAGE, protein stabilization via protein aggregation assays, and survival through single spot dilution and colony-counting methods under various stress conditions. The pre-treatment of short stress dosage showed endured overall tolerance of bacterium to lethal conditions, as evidenced by moderately enhanced total soluble intracellular protein content, better protein stabilization, comparatively over-expressed HSPs, and relatively enhanced cell survival. The findings highlighted that cells grown under optimal conditions were more susceptible to lethal environments than stressed cells, with their enhanced tolerance linked to the overexpression of 20 distinct HSPs of 17-91 kD. These insights offer the potential for developing strategies to enhance microbial resilience for various applications including bacterial bioprocessing, bio-remediation, and infectious disease management.

Enhancing Cd and Pb tolerance of Robinia pseudoacacia (black locust) by regulating antioxidant defense system, macroelement uptake and microstructure.

Woody plants had received considerable attention for the phytoremediation of heavy metal-contaminated soil. This study aimed to investigate the changes in antioxidant enzyme activity, macroelement uptake, and microstructure of the woody plant Robinia pseudoacacia (black locust) for the phytoremediation of cadmium (Cd) and lead (Pb) co-contaminated soil based on dynamic sampling. The results showed that black locust demonstrated strong tolerance in Cd and Pb co-contaminated soil. After 30-120 d of cultivation, the activities of superoxide dismutase (SOD), peroxidase (POD), and the macroelement (K and Ca) content in plant leaves significantly declined in response to Cd and Pb. However, after 160 d of cultivation, the antioxidant enzyme activities, chlorophyll, sulfhydryl, and soluble protein contents, as well as Ca and Mg content in plant leaves were returned to normal levels under the 40mg·kg-1 Cd and 1000mg·kg-1 Pb contaminated soil (CdPb3). Meanwhile, K content in plant leaves under the CdPb3 treatment was significantly (P<0.05) increased by 68.9% compared to the control. Cd and Pb was primarily accumulated in black locust roots. Scanning electron microscope (SEM) analysis indicated that the sieve tubes in the roots and stems of plant might block the transport of Cd and Pb. Transmission electron microscope (TEM) analysis indicated that the number and volume of osmiophilic particles in plant leaves were increased and the cell walls were thickened in response to Cd and Pb stress. Path analysis further indicated that the growth of plant was related to macroelements uptake and physiological change (photosynthesis, antioxidant enzyme activity, and chelation). Thus, black locust could effectively regulate the antioxidant defense system, macroelement absorption, and microstructure to enhance plant tolerance to Cd and Pb stress. Moreover, black locust could maintain the normal urease, acid phosphatase, and sucrase activities in the Cd and Pb co-contaminated soil. These findings suggested that black locust could be considered as a useful woody plant for the phytostabilization in Cd and Pb-contaminated soil.

Investigating enhancement of protease and lysozyme combination pretreatment on hydrolysis of sludge organics under humic acid inhibition

This study investigated the impact of humic acid (HA) on enzymatic pretreatment efficiency, focusing on sludge properties and HA molecular structure. The results showed that enzymatic pretreatment alleviates HA inhibition, improving hydrolysis efficiency. In the presence of HA, soluble proteins and polysaccharides in the enzyme-cocktail group reached 27.7 mg/L and 23.9 mg/L, 1.4 and 1.3 times higher than the blank group, respectively. The enzyme-cocktail group also had the highest soluble DNA concentration (19.4 mg/L) and the lowest viable cell proportion (69.3 %), indicating effective cell lysis. Enzyme-cocktail pretreatment reduced electrostatic repulsion, enhancing the mobility of extracellular organics. Enzyme interactions with HA released internal hydrolases and decreased amide groups on the HA surface, increasing the availability of biodegradable substrates. Overall, enzymatic pretreatment proves effective in mitigating HA-induced inhibition, thereby improving sludge biodegradation and enhancing carbon recovery in anaerobic fermentation.

Biochemical and structural insights into pinoresinol hydroxylase from Pseudomonas sp.

The vanillyl alcohol oxidase/p-cresol methylhydroxylase (VAO/PCMH) flavoprotein family comprises a broad spectrum of enzymes capable of catalyzing the oxidative bioconversions of various substrates. Among them, pinoresinol hydroxylase (PinH) from the 4-alkylphenol oxidizing subgroup initiates the oxidative degradation of (+)-pinoresinol, a lignan important for both lignin structure and plant defense. In this study, we present a detailed biochemical and structural characterization of PinH from Pseudomonas sp., with focus on its substrate specificity and product formation. PinH was expressed in E. coli and purified as FAD-containing, soluble protein. The flavoenzyme catalyzes the hydroxylation of both (+)-pinoresinol and eugenol. Structural analysis reveals its dimeric form, non-covalent flavin binding, and a large active site. AlphaFold models of the PinH-cytochrome complex demonstrate cytochrome's dual role in electron transfer and modulating PinH’s conformation. A distinctive feature of PinH is a large cavity that hosts its multi-ring (+)-pinoresinol substrate. The capability of converting bulky lignans is particularly attractive for biotechnological applications aimed at producing high-value compounds from phenolic precursors. These insights expand our knowledge on the structure and mechanism of the VAO/PCMH flavoenzyme family members.

Yeast extract elicitation enhances growth and metabolite production in Limonium algarvense callus cultures

Limonium algarvense Erben, a medicinal halophyte, holds significant pharmacological promise due to its rich bioactive compound repertoire. This study aimed to establish robust callus cultures as a sustainable, in vitro model for studying the plant's metabolic responses, particularly focusing on synthesising and accumulating primary and secondary metabolites under various elicitation treatments. Callus cultures were initiated from leaf explants on Murashige and Skoog's medium supplemented with 1 mg/L picloram for 4 weeks. Afterwards, callus cultures were subjected to two elicitor treatments, including salicylic acid–SA and yeast extract–YE at 50 and 100 mg/L for four weeks. Water extracts were assessed for their shifts in primary (total soluble sugars and proteins, and proline), and secondary metabolism (total phenolics, flavonoids, and condensed tannins). In addition, a detailed metabolic profiling was conducted using high-performance liquid chromatography with electrospray ionisation mass spectrometry (HPLC–ESI–MS/MS). Elicitation induced significant shifts in the metabolite synthesis of elicited cultures compared to controls. While YE50 markedly increased the callus yield, the total levels of phenolics, flavonoids condensed tannins and total soluble proteins, the SA50 led to the highest increase in proline content. Metabolomic analysis identified 10 metabolites, including 4-hydroxyphenyllactic acid, hydroxybenzoic acid, riboflavin (Vitamin B2), and dihydroferulic acid methyl ester 4-O-sulfate, that were increased in the YE50 elicitation treatment. This suggests that elicitation can effectively enhance the biosynthesis of primary and secondary metabolites in L. algarvense callus cultures, offering great potential for nutritional and medicinal applications.

Integrating physiological, metabolome and transcriptome revealed the response of maize seeds to combined cold and high soil moisture stresses.

Combined cold and high moisture stress (CHS) is a prevalent abiotic stress during maize sowing in northeast China, severely affecting the growth of seedlings and seed germination. However, the mechanism underlying seed growth responses to CHS remains unclear. We used Jidan441 (JD441, CHS-resistant) and Jidan558 (JD558, CHS-sensitive) as experimental materials. Treatments of 5-day cold (4°C, CS), high moisture (25%, gravimetric water content, HH), and CHS were initiated at sowing, followed by a return to normal growth conditions (20°C during light/ 15°C during dark, 15%) at 7 days after sowing (DAS). CS, HH, and CHS decreased seed root length and surface area. The reduction in root length and surface area in JD441 due to CHS was less severe than in JD558. We found that the difference between CHS and control in JD441was less than that in JD558 at transcriptional and metabolic levels at 7 DAS. After CHS removal, JD441 exhibited a greater increase in α-amylase activity and antioxidant content than JD558, which facilitated starch decomposition and the rapid removal of O2 - and H2O2 in seeds. The rapid recovery of soluble sugar and soluble protein in JD441 helped maintain osmotic balance. Amino acids and genes related to amino acid metabolism were upregulated in response to combined stress in JD441, whereas they were downregulated in JD558. In conclusion, the stress tolerance of JD441 was attributed to its efficient recovery ability from CHS. This study provides a scientific foundation for exploring seed stress tolerance pathways and developing cold and high-moisture-tolerant hybrids.

Effect of gibberellic acid treatment and cold stratification on breaking combined (physiological + mechanical) dormancy and germination in Sassafras tzumu (Hemsl.) Hemsl seeds

Sassafras tzumu (Hemsl.) Hemsl., known as Chinese sassafras, is an exclusive species of tree found in China, belonging to the Sassafras genus in the Lauraceae family. Its primary mode of propagation is through seeds, which possesses a profound dormancy and irregular germination. In its natural environment, it takes approximately 2-3 years for the seeds to emerge from the ground and begin germinating. Consequently, a rapid and effective method for breaking dormancy would greatly benefitsbn edjhn the production of S. tzumu seedlings. The objective of this investigation was to identify the optimal treatment for breaking dormancy. The S. tzumu seeds were subjected to gibberellic acid (GA3) treatment and cold stratification. The findings indicated that the dormancy type of Sassafras tzumu seeds was combined (physiological + mechanical) dormancy. After 6 months of cold stratification, the germination percentage of seeds treated with various concentrations of GA3 (200mg/L, 500mg/L, and 1000mg/L) all exceeded 50%. Among these treatments, the most effective method was the application of 200mg/L GA3 in combination with 5 months of cold stratification, resulting in a germination percentage of 55%. During the process of cold stratification, soluble proteins, soluble sugars, starch, and fat were broken down into smaller molecules to support embryo growth. Interestingly, the content of soluble sugar exhibited an upward-downward trend, while the content of soluble protein displayed a downward-upward trend. The release of dormancy in S. tzumu seeds was jointly regulated by various endogenous hormones. The content of GA3 demonstrated an upward-downward trend, while the ABA content exhibited a continuous decline with a brief upward trend. In contrast, the IAA content displayed a continuous upward trend, and the ZR content showed no significant change. The ratios of GA3/ABA, IAA/ABA, and ZR/ABA all displayed an upward trend.

Physicochemical Properties of Quinoa (Chenopodium quinoa Willd.) Protein Isolated from Black, Q12, and Titicaca Seed

Background: Quinoa was cultivated in different parts of the world because of the water crisis. On the other view, proteins from plant sources have attracted significant interest. One of the sustainable protein sources is quinoa protein. Objective: The aim of this study was to isolate quinoa protein and determine physicochemical properties for its use in the food industry. Methods: Quinoa Protein Isolates (QPIs) were separated from Quinoa Seed (QS) varieties (Black- -QS, Q12-QS, and Titicaca-QS). The particle size, FTIR, SEM, emulsion activity and stability, protein solubility, and gelation properties were assessed. Results: The Q12-QPI had the highest average particle size, 945 μm. The Black-QPI and Titicaca (T)-QPI had a higher protein content (87.32 ± 1.93, 87.84 ± 1.62% w/w), respectively, and a more condensed structure. The surface morphology of Black-QPI and T-QPI showed regular flat and compact surfaces with some small aggregates. Black-QPI had the most negative zeta potential charge (-38.8 ± 0.03) and stability among the TQPIs. Emulsion capacity was equal among the samples, but emulsion stability was the greatest value (34.48 ± 8.1) in T-QPI. The protein solubility ratio was 70.72, 70.0, and 69.27% in Q12-QPI, T-QPI, and Black-QPI, respectively. The higher elastic performance of Q12-QPI and T-QPI than Black-QPI was seen during the heating steps in the gelation stage. Conclusion: The suitable nutritional and functional resources of Titicaca quinoa protein make it an appropriate candidate to use as a safe food additive.

Enzyme-Assisted Extraction of Proteins from Cauliflower and Broccoli Waste Leaves

This research presents a novel approach to protein extraction from cauliflower (CL) and broccoli (BL) waste leaves by using enzymatic pretreatment, demonstrating its effectiveness on protein yield. Enzymatic pretreatment (pH 4.5, 10 h, t = 35 °C), was performed using commercial enzyme preparations, Viscozyme® L and Vinozyme®, in three different enzyme-to-substrate ratios E/S (0.2%, 2.5%, and 4.8%) of each enzyme. Leaf proteins (LPs) were obtained with alkaline extraction at pH 10–11 and their isoelectric precipitation at pH 4, following the control sample (extraction without enzymes). Protein yield (%), which was used as a parameter to monitor enzymatic efficiency, demonstrated a direct correlation with the enzyme-to-substrate (E/S) ratio. The highest protein yields were obtained at an enzyme concentration of 4.8% for both cellulolytic and pectolytic enzyme preparations, yielding 14.90 ± 0.12% for CL and 29.88 ± 0.86% for BL. The obtained proteins were characterized by FTIR spectroscopy and SDS-PAGE electrophoresis, and these methods confirmed the enzymatic efficiency of protein isolation. Isolated LPs showed high protein content for CLP 4.8% (77.27 ± 0.14%) and BPL 4.8% (84.66 ± 0.51%), and an increase in total amino acids, while the content of essential amino acids was over 40%. Protein solubility was assessed, revealing significant improvements (p &lt; 0.05) in LPs derived from CL and BL at the highest E/S ratio of 4.8%, compared to the control sample C0%. Specifically, the solubility of CLP reached 29.4 mg/mL at pH 11, while BLP achieved 36.4 mg/mL at pH 10. As a result, these leaf proteins not only meet nutritional demands but also open innovative avenues of research in food science and biotechnology.

High pressure processing at different hydration levels as a tool to enhance rice bran stability and techno-functionality.

High-pressure processing (HPP) enhances food safety and shelf life by inactivating microorganisms and preserving food quality, yet its effectiveness in low-humidity environments has not been evaluated. This study investigated the effects of HPP at 500MPa for 15min across varying hydration levels (15, 30, 60, 77%) on rice bran (RB), aiming to identify microbial effectiveness, besides techno-functional and physicochemical properties. HPP effectively reduced mesophilic bacteria, molds and yeast of RB at>15% hydration level, achieving reductions of up to 4 logarithmic cycles in the latter, nearing the detection limit of the method. However, it did not significantly impact spore inactivation. HPP treatment of≥30% hydrated RB induced particles aggregation and a honeycomb formation. The interaction between hydration and HPP treatment significantly affected the distribution of total dietary fibers, with an increase in soluble dietary fiber from 8.73g/100g to 11.03g/100g after HPP treatment at 15% hydration level. Protein solubility was enhanced by hydration (15, 30 and 60%), and peroxide values decreased after HPP treatment at low hydration (≤30%) but increased when applied to high hydrated (>30%) RB. Emulsifying activity decreased upon HPP treatment of highly hydrated RB (≥60%), but more stable emulsions were achieved after HPP, regardless of the hydration level. Therefore, this study highlights the potential of HPP as a sustainable approach to enhance the utilization of rice bran in food applications, addressing existing knowledge gaps regarding its processing under different moisture conditions.

Berberine enhances autophagic flux to alleviate ischemic neuronal injury by facilitating N-ethylmaleimide-sensitive factor-mediated fusion of autophagosomes with lysosomes.

Our previous study demonstrated that Berberine (BBR) significantly enhances autophagic flux, alleviating ischemic neuronal injury by restoring autolysosomal function, but how BBR augmented autolysosomal functions remained elusive. N-ethyl-maleimide sensitive factor (NSF) is considered as a major ATPase to reactivate soluble NSF attachment protein receptors (SNAREs), which directly mediate autophagosome-lysosome fusion. However, NSF was dramatically inactivated by ischemia to hamper membrane-membrane fusion, leading to autophagic/lysosomal dysfunction in neurons. This study was to investigate whether BBR-ameliorated autophagic flux was exerted by reinforcing NSF activity, which subsequently boosted autophagosome-lysosome fusion in ischemic neurons. Rat model of ischemic stroke and neuronal ischemia model of HT22 cells were prepared by middle cerebral artery occlusion (MCAO) and oxygen-glucose deprivation (OGD), respectively. BBR was intraperitoneally administrated with 100 mg/Kg/d for 3 days before MCAO and was treated with 90 μM in HT22 neurons for 12 h, respectively. The results illustrated that NSF activity was markedly reinforced to facilitate autophagosome-lysosome fusion in penumbral cells and OGD HT22 neurons by BBR treatment. Consequently, the ischemia-created autophagic/lysosomal dysfunction was greatly restored to alleviate ischemic injury. Thereafter, NSF activity in OGD HT22 neurons was altered by transfection with NSF-overexpressing lentiviruses and siRNA-mediated knockdown, respectively. The data showed that BBR-enhanced autophagic flux and it-induced neuroprotection were greatly counteracted by NSF knockdown. By contrast, NSF overexpression synergistically boosted autophagosome-lysosome fusion and further attenuated neuronal death upon BBR treatment. Therefore, our study indicates that BBR-conferred neuroprotection against ischemic stroke is induced through facilitating autophagosome-lysosome fusion, by which enhancing autophagic flux in ischemic neurons.

Solubilization effects of egg yolk granules induced by weakly alkaline treatment.

Egg yolk granules (EYGs) are water-insoluble components of egg yolk that are rich in protein and phospholipids. Improving the solubility of EYGs is crucial for their development and utilization. The solubilization of EYGs in a weak alkaline environment (pH7.5-8.5) was investigated through physicochemical properties, microstructure analysis, and metabolomics. The results reveal a significant reduction in the average particle size of EYGs, ranging from 70.49 to 91.28nm in weakly alkaline conditions. Microstructure and spectroscopic analyses indicate conformational changes in EYGs under these conditions, facilitating the solubilization of proteins, calcium, phosphorus and other components. Furthermore, metabolomics analyses confirms that depolymerization of EYGs promotes the release of glycerophospholipids. These findings underscore the beneficial effects of weakly alkaline conditions on the solubility of EYGs.

Effect of rutin on the structural and functional properties of ovalbumin.

Phenolic substances affect protein functionality. This study aimed to examine how rutin influences the gel properties, antioxidant activity, and structure of ovalbumin (OVA). Increasing rutin concentration enhanced the gel hardness of OVA but reduced soluble protein content with no significant effect on water retention. At 0.25 % rutin concentration, the gel hardness of OVA increased from 109.33 g to 292.60 g, while soluble protein content decreased from 1.08 mg/mL to 0.97 mg/mL. Rutin modification significantly increased the storage and loss moduli of OVA gel, making its structure more compact. At 0.25 % rutin, antioxidant analysis showed increases in the DPPH radical scavenging rate (127 %), ABTS radical scavenging rate (112 %), hydroxyl radical scavenging rate (4167 %), and reducing power (101 %) of OVA. Fluorescence spectroscopy, surface hydrophobicity, free sulfhydryl content, and circular dichroism spectra revealed that higher rutin concentrations reduced fluorescence intensity and surface hydrophobicity while increasing the free sulfhydryl content of OVA. The α-helix content of OVA decreased, while β-sheet content increased. In addition, it was confirmed that OVA and rutin were bound by hydrophobic interaction. The quenching mechanism was static quenching. Rutin alters the structure and functional properties of OVA, providing a theoretical foundation for developing antioxidant and high-gel OVA variants.