Protein Fractions Research Articles

Pea and soy protein isolate fractal gels: The role of protein composition, structure and solubility on their gelation behaviour

The gelation behaviour of two different pea protein isolates and one soy protein isolate were investigated with a focus on the role of the protein properties. Protein solubility was the lowest in pH 3 citrate-phosphate buffer (<10% w/w), increased in pH 7.4 phosphate-buffered saline (12–21% w/w), and was the highest in pH 7.6 MilliQ water (∼20–40% w/w). Heat-induced gelation conditions for the protein sources were sensitive to both the soluble and the insoluble fractions as obtained during extraction. At low protein concentrations (≤5% w/v), the proteins started to lose their viscoelastic behaviour and exhibited predominantly viscous properties. Fitting of the fractional Kelvin-Voigt model to the frequency sweeps showed an increase in the fractal gel strength with increasing protein concentration. Secondary structures of the soluble species showed mostly unordered proteins, suggesting that the proteins were denatured during the commercial extraction process although gelation has to date been suggested to be highly dependent on the denaturation of soluble proteins. Synchrotron Radiation Circular Dichroism measurements of the insoluble proteins showed a significant amount of ordered protein structures. SEM imaging of the gels also suggested a new gelation pathway in which insoluble proteins act as dispersed fillers within a continuous matrix of soluble proteins. This research elucidated the role of different protein fractions, globulins and albumins, and their secondary structure in the formation of a gel network and how this affects their viscoelastic behaviour.

Relationship between soybean protein composition and breaking load of heat-induced gel prepared from acid-precipitated soybean flour

To understand the effective soy protein composition for realizing appropriate meat substitute, we evaluated the contribution of four protein fractions separated from soybeans, two types of globulins and two types of lipophilic proteins, to textural properties of heat-induced gels prepared from an acid-precipitate of soybean flour (AP-SF). When we added these fractions sduring the preparation of AP-SF gels, which increased the protein content by 10%, the gel with 7S globulin displayed the highest breaking load (417.7 gf). With respect to combinatorial effects, the AP-SF gel that added with 5% 7S and 5% 11S globulin reached an even higher value (460.2 gf) than the value described above (417.7gf).To further support the contribution of fractions, we investigated the correlation between the content of these four protein fractions in soybean flour from different cultivars and the breaking load of the heat-induced gels. The breaking load shows a stronger correlation with the total amount of 7S and 11S (r = 0.86) than with 7S alone (r = 0.77), 11S alone (r = 0.70), or the total amount of acid-precipitated proteins (r = 0.80). Thorough this research, we obtained new findings suggesting a synergistic effect between both globulins.

Wall-mounted ultrafine hollow fiber membrane module for purification of domestic surface water

The World Health Organization (WHO) estimated that around 844 million people worldwide lack safe drinking water, including 159 million who are dependent only on surface water. Investing in a practical, sustainable, and cost-effective treatment process is indispensable to alleviating the effect of water shortage. Besides water treatment, ultrafiltration (UF) membranes are broadly used in protein fractionation and clarification of fruit juices. The hollow fiber (HF) membrane configuration has a much larger surface area per unit volume, resulting in higher productivity than the flat-sheet membrane. A spinneret was developed with a provision for altering the fiber's inner diameter from 1000 µm to 150 µm and outer dimension from 1500 µm to 350 µm, respectively, using an inbuilt detachable needle arrangement. The ultrathin hollow fiber produced by the indigenously designed spinnerets has a high surface area and packing density, which makes it useful for surface water treatment. A wall-mounted module was designed for direct connection to the tap to obtain a purified water flow of 30–40 liter per hour (LPH) for domestic use, operated with or without electricity. The fabricated ultrafine HF module (ID: 150–200 µm; OD: 350–450 µm) removes complete bacterial contamination and turbidity up to 95–100 %, making the quality of processed water potable. The developed membrane module is durable due to its large membrane area per unit volume, enhancing productivity, high packing density, good fouling resistance, easy handling, and self-supporting structure for backwashing. The present research article paves the way forward to investing in a practical, sustainable, and economical treatment process that is indispensable to alleviating the effect of water shortage.

Optimizing Feed Formulation Strategies for Attaining Optimal Nutritional Balance in High-Performing Dairy Goats in Intensive Farming Production Systems

Abstract Genetic factors play a pivotal role in shaping feed formulation strategies for dairy goats, influencing milk production, composition, and overall productivity through the modulation of genetic parameters and polymorphisms associated with milk protein fractions and yield. Moreover, environmental challenges, including heat stress and feed availability fluctuations, pose significant hurdles in achieving optimal nutrition and performance, necessitating precision livestock farming and the utilization of local by-products to mitigate their impact on herd health and productivity. In addition, nutritional interventions, such as supplement utilization and alternative feed incorporation, offer promising avenues for addressing specific deficiencies and enhancing overall performance, from fatty acid supplementation to herbal feed additives, while balancing rations to meet evolving nutritional needs and ensuring long-term sustainability remains paramount for optimizing herd nutrition, health, and performance in intensive dairy goat production systems. The aim of this study was to explore the multifaceted influences of genetic factors, environmental challenges, and nutritional interventions on feed formulation strategies and overall herd nutrition, health, and performance in intensive dairy goat production systems.

Characteristics of isatuximab-derived interference in serum protein electrophoresis and immunofixation, and an absence of sustained in vivo interference due to belantamab mafodotin and denosumab

ObjectivesSome therapeutic monoclonal antibodies, like daratumumab and elotuzumab, produce interfering monoclonal bands on serum protein electrophoresis (SPEP) and immunofixation electrophoresis (IFE). Whether other common therapeutic antibodies also produce interference has not been systematically evaluated. Design and methodsSPEP/IFE from patients receiving isatuximab (48 patients), belantamab mafodotin (BM; 41), and denosumab (41) were retrospectively reviewed for therapeutic antibody interference. Cases exhibiting isatuximab interference were quantified and the maximum duration of isatuximab effect was evaluated. To characterize band position, neat human serum was spiked with BM or denosumab at supratherapeutic concentrations. Band migration patterns were compared on SPEP and IFE, with band position expressed relative to other constant protein fractions. ResultsIsatuximab-induced IFE interference was common (81.3 % of evaluated patients) with a maximum observed duration of 8 weeks. 10.4 % of isatuximab patients had IgG kappa monoclonal gammopathies that co-migrated with the drug; this subset could benefit from HYDRASHIFT 2/4 isatuximab testing. 8.3 % of IFE cases were negative for an isatuximab band but showed large, endogenous M-spikes migrating elsewhere. All patients in this group expired within 1 year of this finding. We hypothesize that an inability to detect isatuximab in this setting corresponds to a large residual myeloma burden that reduces isatuximab serum concentration. This observation may serve as a negative prognostic factor. Spiking studies demonstrated that BM and denosumab produce interference in vitro, but sustained interference was not observed in >40 treated patients. ConclusionsTherapeutic antibody interference in patients receiving isatuximab is common, and can persist for at least 8 weeks after administration. >10 % of patients receiving isatuximab may benefit from HYDRASHIFT testing post-therapy. In contrast, BM and denosumab fail to produce sustained interference in treated patients.

Streptococcus thermophilus: A Source of Postbiotics Displaying Anti-Inflammatory Effects in THP 1 Macrophages.

In addition to traditional use in fermented dairy products, S. thermophilus also exhibits anti-inflammatory properties both in live and heat-inactivated form. Recent studies have highlighted that some hydrolysates from surface proteins of S. thermophilus could be responsible partially for overall anti-inflammatory activity of this bacterium. It was hypothesized that anti-inflammatory activity could also be attributed to peptides resulting from the digestion of intracellular proteins of S. thermophilus. Therefore, total intracellular proteins (TIP) from two phenotypically different strains, LMD-9 and CNRZ-21N, were recovered by sonication followed by ammonium sulphate precipitation. The molecular masses of the TIP of both strains were very close to each other as observed by SDS-PAGE. The TIP were fractionated by size exclusion fast protein liquid chromatography to obtain a 3-10 kDa intracellular protein (IP) fraction, which was then hydrolysed with pancreatic enzyme preparation, Corolase PP. The hydrolysed IP fraction from each strain exhibited anti-inflammatory activity by modulating pro-inflammatory mediators, particularly IL-1β in LPS-stimulated THP-1 macrophages. However, a decrease in IL-8 secretion was only observed with hydrolysed IP fraction from CNRZ-21N, indicating that strain could be an important parameter in obtaining active hydrolysates. Results showed that peptides from the 3-10 kDa IP fraction of S. thermophilus could therefore be considered as postbiotics with potential beneficial effects on human health. Thus, it can be used as a promising bioactive ingredient for the development of functional foods to prevent low-grade inflammation.

QUALITY OF MARE'S MILK AND KOUMISS

Nowadays mare's milk and koumiss are in great demand in Kazakhstan. Therefore, the study of its qualitative indicators is relevant.In the article the physical and chemical parameters and vitamin content (B and C groups) of mare's milk and koumiss samples taken from peasant farms of Almaty region in the fall-winter period are studied in a comparative form.The study showed that mare's milk samples had no significant difference in total protein mass fraction in the fall and winter. In mass fraction terms of fat in the sample obtained in the fall the fat content was 2.18% higher than in the sample obtained in winter. And the mass fraction of lactose in the sample obtained in the fall was 0.14% higher than in the sample obtained in winter.According to the study results of obtained koumiss samples’ physical and chemical parameters in the fall and winter periods, the mass fraction of protein was 1.86-1.89%. Fat content of koumiss proportion prepared in the fall period was 1.84%, in the winter period-2.19%, and the average mass fraction of lactose was from 3.64 to 3.98%.The content of water-soluble vitamins B and C in samples of mare's milk and koumiss was investigated. According to the study results the content of vitamins B1, B2, B3, B5 B6 and C in the composition of mare's milk and koumiss obtained in the fall and winter periods was established.According to the general research results samples of mare's milk and koumiss products obtained from peasant farms of Almaty region have proven high quality, nutritious in food value and easily assimilated by human body.

Tunisian Silybum Species: Important Sources of Polyphenols, Organic Acids, Minerals, and Proteins across Various Plant Organs.

Silybum marianum and Silybum eburneum are wild edible Mediterranean plants used in the human diet. This study presents the initial findings on the phytochemical characterization of Tunisian S. marianum and S. eburneum organs. It examined their mineral, sugar, organic acid, polyphenolic, and seed storage protein contents, as well as their antioxidant potential. In S. marianum, stems had high sodium and potassium contents, while the immature and mature seeds were rich in calcium and magnesium. However, S. eburneum had high potassium levels in stems and high sodium and calcium levels in the flowers. S. marianum showed substantial fructose variation among its organs. Conversely, S. eburneum exhibited significant heterogeneity in glucose, sucrose, and maltose levels across its organs, with maltose exclusively detected in the immature seeds. A notable organ-dependent distribution of organic acids was observed among the two species. Higher levels of phenolic contents were detected in both mature and immature seeds in both species compared to the other plant parts. The seeds possessed higher antioxidant activities than other plant organs. In both S. marianum and S. eburneum seeds, albumins and globulins were the predominant protein fractions. This study brings evidence supporting the important potential of Silybum organs as sources of nutrients with antioxidant properties for producing functional food.

Phylogeny and Expansion of Serine/Threonine Kinases in Phagocytotic Bacteria in the Phylum Planctomycetota.

The recently isolated bacterium "Candidatus Uabimicrobium amorphum" is the only known prokaryote that can engulf other bacterial cells. Its proteome contains a high fraction of proteins involved in signal transduction systems, which is a feature normally associated with multicellularity in eukaryotes. Here, we present a protein-based phylogeny which shows that "Ca. Uabimicrobium amorphum" represents an early diverging lineage that clusters with the Saltatorellus clade within the phylum Planctomycetota. A gene flux analysis indicated a gain of 126 protein families for signal transduction functions in "Ca. Uabimicrobium amorphum", of which 66 families contained eukaryotic-like Serine/Threonine kinases with Pkinase domains. In total, we predicted 525 functional Serine/Threonine kinases in "Ca. Uabimicrobium amorphum", which represent 8% of the proteome and is the highest fraction of Serine/Threonine kinases in a bacterial proteome. The majority of Serine/Threonine kinases in this species are membrane proteins and 30% contain long, tandem arrays of WD40 or TPR domains. The pKinase domain was predicted to be located in the cytoplasm, while the WD40 and TPR domains were predicted to be located in the periplasm. Such domain combinations were also identified in the Serine/Threonine kinases of other species in the Planctomycetota, although in much lower abundances. A phylogenetic analysis of the Serine/Threonine kinases in the Planctomycetota inferred from the Pkinase domain alone provided support for lineage-specific expansions of the Serine/Threonine kinases in "Ca. Uabimicrobium amorphum". The results imply that expansions of eukaryotic-like signal transduction systems are not restricted to multicellular organisms, but have occurred in parallel in prokaryotes with predatory lifestyles and phagocytotic-like behaviors.

Manipulation of the post-flowering source/sink ratio differentially affects protein composition and gluten quality in contrasting bread wheat genotypes

Studies of the impact of assimilate availability during grain filling on protein composition and gluten quality of bread wheat are scarce. Therefore, this work aimed to analyze the responses of wet gluten percentage, SDS sedimentation index, and protein composition to changes in the source/sink (S/S) ratio (post-flowering growth per grain) in six Argentinean wheat genotypes contrasting in baking aptitude (three quality groups), as well as to evaluate its interaction with early nitrogen-sulfur fertilization. For two years, rainfed field trials were carried out in the Humid Pampas in Southern South America, applying manipulative S/S treatments (shading, defoliation, thinning, and trimming). Gluten percentage showed a negative linear trend against the increase in S/S ratio with differences among genotypes even within the same quality group (slope range: −0.0248 to −0.6566). The associations between the main protein fractions (i.e., GLI/GLU, HMW/LMW) with the S/S ratio were not significant, while the proportions of specific high molecular weight glutenin subunits were relevant, such as Glu-A1x/HMW (slope range: −0.00027 to −0.00147) and Glu-D1y/HMW (0.00029–0.00139). Early nitrogen-sulfur fertilization increased the influence of the S/S ratio on gluten percentage while stabilizing the sedimentation index. These results apport another approach, based on S/S ratio, to understanding the determination of grain quality under changing contexts.

Exploring the role of production and release of proteins for microbial interactions in kombucha

Driving kombucha fermentation through microbial consortia remains based on scarce scientific elements, especially regarding the consumption of nitrogenous nutrients. This study aimed at analyzing quantitatively proteins and free amino nitrogen (FAN) concentrations, and qualitatively the soluble proteins profiles during a 12-day production process. The effect of microbial interactions was evaluated by comparing the growth of previously isolated major microorganisms in monoculture and cocultures to original kombucha. Protein and FAN relative concentrations kinetics depended on microbial compositions. These parameters were associated to less variations and a positive effect on total microbial population in original kombucha, underlining a complex yet better management of the nutrients. Analysis of extracellular proteins by electrophoresis showed the presence of a microbial 15 kDa fraction common to all cultures. In contrast, a 21 kDa protein fraction was associated with the presence of Acetobacter indonesiensis. Exploratory proteomics analysis revealed the presence of characteristic extracellular moonlighting proteins, originating from this acetic bacteria. Production of extracellular vesicles was successfully confirmed with the isolation and observation of vesicles in A. indonesiensis culture, as well as in kombucha. The results characterize the central role of microbial interactions on FAN consumption and open new perspectives on the function of the bacterial vesicles observed.

Amino acid composition of pollen &lt;i&gt;Pinus sylvestris&lt;/i&gt; L. and &lt;i&gt;Pinus sibirica&lt;/i&gt; Du Tour growing in the Baikal region

The purpose of the study was to determine the qualitative and quantitative amino acid composition of pollen Pinus sylvestris L. and Pinus sibirica Du Tour. Pine pollen was collected in June 2021 at natural sites on the southeastern coast of Lake Baikal. The mass fraction of crude protein was determined by the Kjeldahl method; the protein composition and individual amino acid content were determined by high-performance liquid chromatography with post-column modification of ninhydrin on an LA8080 automatic analyser (Hitachi, Japan). The crude protein content of the pollen was 14.38–15.94%. Pine pollen protein is shown to contain 17 amino acids, including 9 essential amino acids: valine, isoleucine, leucine, threonine, methionine, phenylalanine, lysine, histidine, and arginine. The content of the sum of amino acids was 141.4–156.5 mg/g, including essential amino acids 45.9-48.4%. The following amino acids are dominant in Pinus sylvestris and Pinus sibirica pollens (mg/g): monoaminodicarboxylic acids – glutamic (21.3–24.2) and aspartic (13.0–14.2), diaminocarboxylic acid arginine (17.0–17.4) and heterocyclic amino acid proline (14.7–16.2). The obtained results can be useful in the development of drugs and biologically active additives based on pollen Pinus sylvestris and Pinus sibirica, which, due to the presence of the above amino acids, have a nootropic, immunomodulatory, cardiac stimulating, and detoxifying effect.

Физико-химические показатели гомогената трутневого расплода, адсорбированного на безлактозных адсорбентах в процессе хранения

Abstract. Drone brood is a product that exhibits high biological activity, so its use can be recommended to a wide range of people as an adaptogenic, actoprotective and antioxidant agent. The high concentration of biologically active substances, such as decenoic acids, unsaturated fatty acids, sulfhydryl groups, sugars, hormones and hormone-like compounds makes drone brood a product very sensitive to environmental factors, which necessitates its stabilization. To stabilize drone brood homogenate, in a number of countries, a lactose-glucose adsorbent is usually used. The relevance of the work is due to the widespread prevalence of lactase deficiency among the population, the prevalence of which among the Eastern Slavs reaches 16–18 %, and among some peoples of Asia, Australia and America exceeds 80 %, which limits the use of adsorbed drone brood using a classical adsorbent containing lactose. The purpose of the work is to develop an innovative product based on drone brood that does not contain lactose as an adsorbent, and to trace the dynamics of the preservation of its biologically active components during storage. Potato starch was used as a lactose substitute in the adsorbent. This product can be recommended to a wider range of consumers. The work used methods for determining the physicochemical parameters of products, such as humidity, oxidation index, hydrogen index, mass fraction of decenoic acids, mass fraction of crude protein, free acidity, acid number, iodine number. The scientific novelty lies in the development of formulations for new products based on drone brood that do not contain lactose and the determination of the optimal product among them in terms of technological and physicochemical parameters. As a result of the studies, it was found that an adsorbent consisting of 80 parts of starch and 20 parts of glucose provides optimal product performance at a ratio of adsorbent to drone brood homogenate of 3 : 1.

Development of technology for obtaining an enriched fermented milk product based on the results of studies of the composition of the initial and final materials

The article presents the results of research on the study of the chemical, mineral and amino acid composition of the initial and final materials of the production technology of a fermented milk product enriched with an extract of vegetable raw materials. As a sample of the initial product, the optimal composition of a combination of two types of dairy raw materials, 15% mare's milk and 85% cow's milk, obtained during experiments, was selected. The final product is a fermented milk product obtained from combined dairy raw materials. It was found that in the combined dairy raw materials, the mass fraction of protein is 3.86%, the mass fraction of fat is 4.66%, the mass fraction of carbohydrates is 4.95%, in the fermented milk product, the mass fraction of protein is 4.04%, the mass fraction of fat is 4.85%, the mass fraction of carbohydrates is 6.05%. The energy value of the fermented milk product, in kcal -82.5, of the combined feedstock -75.94 kcal. The amino acid composition in the studied samples showed the presence of such components as – arginine, lysine, tyrosine, phenylalanine, histidine, leucine+isoleucine, methionine, valine, proline, threonyl, serine, alanine and glycine. The mineral composition of the combined dairy raw materials in weight% is 14.15 calcium, 15.41 potassium, 12.36 phosphorus, 1.38 magnesium and 7.34 sodium. The results of the analyses confirmed the nutritional and biological value of both the initial and final products, the technology being developed for the production of functional dairy products.

Determining the Impact of Genotype × Environment on Oat Protein Isolate Composition Using HPLC and LC-MS Techniques.

The effect of genotype and environment on oat protein composition was analyzed through size exclusion-high-performance liquid chromatography (SE-HPLC) and liquid chromatography-mass spectrometry (LC-MS) to characterize oat protein isolate (OPI) extracted from three genotypes grown at three locations in the Canadian Prairies. SE-HPLC identified four fractions in OPI, including polymeric globulins, avenins, glutelins, and albumins, and smaller proteins. The protein composition was dependent on the environment, rather than the genotype. The proteins identified through LC-MS were grouped into eight categories, including globulins, prolamins/avenins, glutelins, enzymes/albumins, enzyme inhibitors, heat shock proteins, grain softness proteins, and allergenic proteins. Three main globulin protein types were also identified, including the P14812|SSG2-12S seed storage globulin, the Q6UJY8_TRITU-globulin, and the M7ZQM3_TRIUA-Globulin-1 S. Principal component analysis indicated that samples from Manitoba showed a positive association with the M7ZQM3_TRIUA-Globulin-1 S allele and Q6UJY8_TRITU-globulin, while samples from Alberta and Saskatchewan had a negative association with them. The results show that the influence of G × E on oat protein fractions and their relative composition is crucial to understanding genotypes' behavior in response to different environments.

Dietary guanidinoacetate reduces spaghetti meat myopathy risk in the breast muscle of broiler chickens

The global demand for white chicken meat along with the increase in the occurrence of growth-related breast muscle myopathies (BMMs) [namely white striping (WS), wooden breast (WB), and spaghetti meat (SM)] highlights the need for solutions that will improve meat quality while maintaining the high productivity of modern broilers. Guanidinoacetate (GAA), a precursor of creatine, is used as a feed additive and has previously shown the potential to affect the quality of breast meat. This study investigated growth performance, meat quality and the risk ratio for the development of BMMs in broilers assigned to two dietary treatments: control (CON) group, fed a commercial basal diet, and supplemented GAA (sGAA) group, receiving the control diet supplemented on top with 0.06% GAA. Growth performance indicators such as BW, daily weight gain, daily feed intake, feed conversion ratio and cumulative feed conversion ratio were recorded on a pen basis. As a trait affecting animal welfare, the occurrence of foot pad dermatitis was also evaluated. At day 43, birds were processed, and breasts were scored for the incidence and severity of BMMs (n = 166 and 165 in CON and sGAA groups, respectively). Quality traits (ultimate pH, colour) and technological properties (i.e., drip and cooking losses, marinade uptake, shear force, and oxidation levels of the lipid and the protein fractions) of breast meat were assessed in both treatments on samples not showing any macroscopic sign of BMMs (n = 20 breast fillets per group). Data of myopathy risk ratio were analysed as the risk for each group to develop WS, WB, and SM myopathies. Our results show that while sGAA and control groups did not differ significantly in growth performance, a remarkably beneficial effect of GAA was observed on the incidence of BMMs with significantly reduced risk of sGAA group to develop SM myopathy. The risk of sGAA group to develop SM was 30% lower compared to CON (P = 0.028). Finally, a significantly lower drip loss was observed in sGAA in comparison with CON (1.78 vs 2.48%, P = 0.020). Together, our results show that the inclusion of 0.06% GAA in feed can improve the water-holding capacity of meat and reduce the risk to develop SM myopathy without compromising the performance of broilers.

Reproduction of the technology for the production of “Rossiysky” cheese in laboratory conditions, depending on the quality of milk

Relevance. Dairy processing enterprises are expanding the range of dairy products with the introduction of production of various cheeses. Small producers have also become active in cheese production. In 2023, the decrease in the price and demand for raw milk had a particularly strong impact on collective (farmer) and personal subsidiary farms. This caused the need for milk processing in rural conditions. Our research is aimed at developing technology for the production of ”Rossiysky” cheese for peasants and farms.Methods. The Kjeldahl method determined the mass fraction of protein, acid method — the mass fraction of fat, calculated — the mass fraction of dry matter and skimmed milk residue, ultrasonic — the mass fractions of milk fat, protein, lactose, milk powder, skimmed milk residue, the amount of added water, mineral salts, density, freezing point, temperature, by the express method — active acidity.Results. The chemical composition and physico-chemical properties of raw milk were studied in winter and spring. It was found that the mass fraction of fat in milk reaches 4.31–4.65%. Mass fraction of protein is 2.92–3.02%. The ratio of milk fat to protein is 1.48–1.54, with the recommended norm being 1.1–1.25. Changes were made to the technological scheme for the production of “Rossiysky” cheese, taking into account the characteristics of the quality of milk and the equipment used in the conditions of peasants and farms. To obtain milk with the required protein to fat ratio, the mixture was normalized. “Rossiysky” cheese was produced from normalized milk. The quality of cheese in terms of organoleptic, physico-chemical and microbiological indicators meets the requirements of regulatory documents.

Unlocking the potential of waxworm (Galleria mellonella) proteins: Extraction, fractionation, and protein quality assessment

Edible insects are increasingly recognized as an alternative and sustainable protein source to meet future protein demands. Owing to its significant protein content (33.43%), the edible insect Waxworm (Galleria mellonella) was characterized in this study for its protein extractability, structural properties, and protein quality. Protein extractability under various pH levels and ionic strengths was evaluated. Subsequently, Osborne fractionation was employed to characterize protein fractions based on their preferential solubilities. The extracted proteins and their respective fractions were visualized by SDS-PAGE, and the most intense bands were excised for proteomics data analysis. Secondary structure, pI, molecular mass, and denaturation temperature were also examined. The quality of waxworm protein was assessed by determining the IVPD, PDCAAS, and amino acid profile. Waxworm proteins exhibited greater stability in alkaline conditions compared to acidic environments. These might also be more sensitive to ionic strength than other alternative proteins, such as those derived from plants. The allergen tropomyosin was identified by SDS-PAGE with an approximate molecular weight of ∼32.5 kDa; however, it can be selectively removed by salt extraction. The secondary structures of waxworm proteins, extracted under various conditions, appeared stable, with α-helix and β-conformation being the predominant conformation. Among the protein fractions, prolamins displayed an α-helix conformation, while albumins, globulins, and glutelins predominantly adopted a β-sheet structure. It was observed that structural features could influence the overall quality of the protein. As for the protein quality, globulins are the protein fraction recommended as these showed the highest EAAI% (924.02), PDCAAS (0.84) and IVPD (76.33%) values.

Alterations in HCN1 expression and distribution during epileptogenesis in rats

BackgroundThe hyperpolarization-activated cyclic nucleotide-gated cation channel (HCN1) is predominantly located in key regions associated with epilepsy, such as the neocortex and hippocampus. Under normal physiological conditions, HCN1 plays a crucial role in the excitatory and inhibitory regulation of neuronal networks. In temporal lobe epilepsy, the expression of HCN1 is decreased in the hippocampi of both animal models and patients. However, whether HCN1 expression changes during epileptogenesis preceding spontaneous seizures remains unclear. ObjectiveThe aim of this study was to determine whether the expression of HCN1 is altered during the epileptic prodromal phase, thereby providing evidence for its role in epileptogenesis. MethodsWe utilized a cobalt wire-induced rat epilepsy model to observe changes in HCN1 during epileptogenesis and epilepsy. Additionally, we also compared HCN1 alterations in epileptogenic tissues between cobalt wire- and pilocarpine-induced epilepsy rat models. Long-term video EEG recordings were used to confirm seizures development. Transcriptional changes, translation, and distribution of HCN1 were assessed using high-throughput transcriptome sequencing, total protein extraction, membrane and cytoplasmic protein fractionation, western blotting, immunohistochemistry, and immunofluorescence techniques. ResultsIn the cobalt wire-induced rat epilepsy model during the epileptogenesis phase, total HCN1 mRNA and protein levels were downregulated. Specifically, the membrane expression of HCN1 was decreased, whereas cytoplasmic HCN1 expression showed no significant change. The distribution of HCN1 in the distal dendrites of neurons decreased. During the epilepsy period, similar HCN1 alterations were observed in the neocortex of rats with cobalt wire-induced epilepsy and hippocampus of rats with lithium pilocarpine-induced epilepsy, including downregulation of mRNA levels, decreased total protein expression, decreased membrane expression, and decreased distal dendrite expression. ConclusionsAlterations in HCN1 expression and distribution are involved in epileptogenesis beyond their association with seizure occurrence. Similarities in HCN1 alterations observed in epileptogenesis-related tissues from different models suggest a shared pathophysiological pathway in epileptogenesis involving HCN1 dysregulation. Therefore, the upregulation of HCN1 expression in neurons, maintenance of the HCN1 membrane, and distal dendrite distribution in neurons may represent promising disease-modifying strategies in epilepsy.

Abstract 4449: Hepatic steatosis induced by bioactive lipids

Abstract Introduction: HCC is the most common form of liver cancer and is the sixth most common cancer globally. Despite the clinical advancement of immune checkpoint inhibitors, median survival continues to be less than twenty months and most patients develop resistance making it one of the deadliest malignancies. The rate of liver cancer continues to rise, which is coupled to the high prevalence of steatosis and steatohepatitis that accounts for about 25% in the general population. Pathologically, 80% of liver cancer occurs in patients with an underlying liver disease that displays liver steatosis. Thus, it is paramount to better characterize the transition from steatosis to HCC. Methods: Utilizing a Pten deleted mouse model (PtenloxP/loxP; Alb-Cre+) that recapitulates HCC progression, protein and lipid fractions were isolated to explore differentially expressed proteins and their correlations with bioactive lipid metabolism. Global untargeted differential proteomics was done with fractionation using a qExactive. Lipid fractions were analyzed using a Sciex 6500 for targeted lipidomic analysis. In addition, genetic knockout hepatocyte cell lines of each AKT isoform were used to explore the involvement of the PI3K/AKT signal. Further, a phosphoproteomic enrichment was performed to discover downstream signaling targets. Results: The PTEN regulated PI3K/AKT signal is induced in 54% of all liver cancers and represents the dominant signaling pathway regulating liver cancer progression. Untargeted differential proteomic analysis of Pten deleted mice livers revealed significant dysregulation in oxidative stress and eicosanoid metabolism among the top enriched disease and biological functions when the phenotype progressed from steatosis to HCC. Analysis of publicly available liver cancer patient samples from the NCI Proteomic Data Commons further shows strong correlation between PTEN protein abundance and the expression of enzymes involved in eicosanoid metabolism. Our analysis using a bio-active lipid multi reaction monitoring panel of Pten deleted mouse livers further validates significant decreases in resolving eicosanoid levels along with increases in proinflammatory eicosanoid precursors as steatosis progress to HCC. These data suggest a potential hepatic AKT-dependency in regulating the shift towards proinflammatory eicosanoids. Primary isolated hepatocytes from mouse livers lacking either AKT1 or AKT2 were analyzed to elucidate AKT’s regulatory role in hepatic eicosanoid metabolism. Proteomic and lipidomic analysis of these hepatocytes supported a unique AKT isoform specific role in the regulation of eicosanoids via potential isoform specific signaling interactions with MAPK. Conclusion: Eicosanoid metabolism dysregulation plays a key role in the progression from steatosis to HCC and appears to be regulated in an AKT isoform specific manner via interactions with MAPK signaling. Citation Format: Mario Alba, Ielyzaveta Slarve, Brandon Ebright, Yiren Zhou, Whitaker Cohn, Yunyi Jia, Elizabeth Elton, Jared Khan, Aditi Datta, Lina He, Qi Tang, Pranav Pammidimukkala, Taojian Tu, Phillip Nguyen, Jonathan Katz, Julian Whitelegge, Stan Louie, Bangyan Stiles. Hepatic steatosis induced by bioactive lipids [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4449.