Protein Extracts Research Articles

Localization of advanced glycation end-products and their receptor in tendinopathic lesions.

This study was designed to investigate the accumulation of advanced glycation end-products (AGEs) and the expression of the receptor of AGEs (RAGE) in tendinopathic tissues. In this study, tendinopathic posterior tibial tendons (PTT) were collected from patients (n=6). Redundant autografts of flexor digitorum longus tendon (FDL; n=3) were used for controls. The control and tendinopathic tendon tissues were used for extraction of proteins for western blot and sectioned for histology and immunohistochemistry. Tendinopathy of the PTT was confirmed histologically by the presentation of disorderly organized collagen fibers, high cellularity and increased vascularity. By immunohistochemistry, heterogeneous accumulation of AGEs was detected on the PTT sections and concentrated in areas, where collagen fibers were disorderly and tangled. In the PTT, roundish tenocytes were also AGEs-positive. In contrast, AGEs were diffuse, lightly stained in the FDL. A greater number of tenocytes within the tendinopathic lesions in the PTT were RAGE positive, compared to the tenocytes in the FDL. Western blot confirmed the expression of AGEs and RAGE in both tendinopathic PTT and control FDL but their band densities were not significantly different. The spatial relation of the accumulated AGEs and RAGE- positive tenocytes within the tendinopathic lesions indicates their involvement in the molecular pathology of tendinopathy.

Lipid-polymer nanoparticles to probe the native-like environment of intramembrane rhomboid protease GlpG and its activity.

Polymers can facilitate detergent-free extraction of membrane proteins into nanodiscs (e.g., SMALPs, DIBMALPs), incorporating both integral membrane proteins as well as co-extracted native membrane lipids. Lipid-only SMALPs and DIBMALPs have been shown to possess a unique property; the ability to exchange lipids through 'collisional lipid mixing'. Here we expand upon this mixing to include protein-containing DIBMALPs, using the rhomboid protease GlpG. Through lipidomic analysis before and after incubation with DMPC or POPC DIBMALPs, we show that lipids are rapidly exchanged between protein and lipid-only DIBMALPs, and can be used to identify bound or associated lipids through 'washing-in' exogenous lipids. Additionally, through the requirement of rhomboid proteases to cleave intramembrane substrates, we show that this mixing can be performed for two protein-containing DIBMALP populations, assessing the native function of intramembrane proteolysis and demonstrating that this mixing has no deleterious effects on protein stability or structure.

Photo‐induced polymerization of styrene‐maleic acid copolymers for the extraction of membrane proteins

AbstractWe report herein the photoinduced electron/energy transfer–reversible addition‐fragmentation chain transfer (PET‐RAFT) polymerization of styrene‐maleic anhydride (SMAnh) using the photocatalyst, zinc tetraphenylporphirine, under a white cool LED light. PET‐RAFT is an easy and convenient polymerization method that does not require deoxygenation unlike other radical polymerizations. Various parameters, for example, the amount of styrene in the feed or the solvent, and their influence on the polymerization were studied. After hydrolysis of the anhydride moieties, the resulting styrene‐maleic acid (SMA) copolymers obtained by PET‐RAFT copolymerization were evaluated for their solubilization efficiency of three different membrane proteins, BmrA and AcrB, overexpressed in Escherichia coli, and A2AR, expressed in insect cells (Sf9). The different copolymers provided similar solubilization rates to the commercially available SMA; however, a highly improved migration behavior on sodium dodecyl‐sulfate polyacrylamide gel electrophoresis was observed which could, facilitate downstream analyses. Overall, we demonstrated that PET‐RAFT is a versatile oxygen tolerant polymerization technique to yield SMA, a suitable polymer for biochemical applications.

Comparative proteomic analysis of retinal hypoxia-ischemia in an acute ocular hypertension model using tandem mass tag-based quantitative proteomics.

The main symptom of acute glaucoma is acute ocular hypertension (AOH), which leads to the death of retinal ganglion cells (RGCs) and permanent loss of vision. However, effective treatments for these conditions are lacking. This study aimed to identify major regulators and overall protein changes involved in AOH-induced RGC death. Proteomic patterns of the retinal protein extracts from the AOH and sham groups were analyzed using mass spectrometry (MS), followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Proteomic analysis revealed 92 proteins in the AOH group compared to the control group; 58 proteins were upregulated and 34 were downregulated. Alterations in fatty acid-binding protein 7 (FABP7) and caveolin-1 (Cav-1), which are related to fatty acid metabolism and ocular inflammatory signaling, were detected using western blotting and biochemical assays. Variations in the expression of galectin-1 (Gal-1), S100 calcium-binding protein A6 (S100a6), and visinin-like protein-1 (VILIP) have been associated with neuronal ischemia. Our investigation demonstrates that neuroinflammation and fatty acid metabolism are involved in retinal impairment following AOH, suggesting a possible treatment approach for acute glaucoma.

A phosphorylation-controlled switch confers cell cycle-dependent protein relocalization.

Tools for acute manipulation of protein localization enable elucidation of spatiotemporally defined functions, but their reliance on exogenous triggers can interfere with cell physiology. This limitation is particularly apparent for studying mitosis, whose highly choreographed events are sensitive to perturbations. Here we exploit the serendipitous discovery of a phosphorylation-controlled, cell cycle-dependent localization change of the adaptor protein PLEKHA5 to develop a system for mitosis-specific protein recruitment to the plasma membrane that requires no exogenous stimulus. Mitosis-enabled anchor-away/recruiter system comprises an engineered, 15 kDa module derived from PLEKHA5 capable of recruiting functional protein cargoes to the plasma membrane during mitosis, either through direct fusion or via GFP-GFP nanobody interaction. Applications of the mitosis-enabled anchor-away/recruiter system include both knock sideways to rapidly extract proteins from their native localizations during mitosis and conditional recruitment of lipid-metabolizing enzymes for mitosis-selective editing of plasma membrane lipid content, without the need for exogenous triggers or perturbative synchronization methods.

Physicochemical Properties, Functionalities, and Antioxidant Activity of Protein Extracts from New Zealand Wild Sea Cucumbers (Australostichopus mollis)

This study investigated the physicochemical properties, functionalities, and antioxidant capacities of protein extracts from wild sea cucumber Australostichopus mollis collected from four distinct locations in New Zealand. Protein was extracted from sea cucumber body walls using trypsin enzymatic extraction, followed by cold acetone precipitation. The amino acid analysis revealed high glycine (189.08 mg/g), glutamic acid (119.45 mg/g), and aspartic acid (91.91 mg/g) concentrations in all samples. The essential amino acid indexes of the protein extracts (62.96, average) were higher than the WHO/FAO standard references, indicating the excellent protein quality of A. mollis. Furthermore, protein extracts from A. mollis demonstrated superior emulsifying activity (202.3–349.5 m2/g average) compared to commercial soy and whey protein isolates under all tested pH conditions, and enhanced foaming capacity (109.9–126.4%) and stability (52.7–72%) in neutral and acidic conditions. The extracts also exhibited good solubility, exceeding 70% across pH 3–11. Antioxidant capacities (ABTS and DPPH free radical scavenging activity and ferric reducing antioxidant power) were identified in A. mollis protein extracts for the first time, with clear variations observed among different locations. These findings elucidate the advantageous functional properties of protein extracts from wild New Zealand A. mollis and highlight their potential application as high-quality antioxidant food ingredients.

Characterisation, antibacterial activity, and prebiotic potential of dried Hermetia illucens L. larvae and of their fractions

Abstract In this study, black soldier fly (BSF) dried larvae were fractionated into their defatted, lipid, protein, and chitin-rich fractions. The samples were then characterised in terms of proximate composition, analysis of amino acids, analysis of the molecular weight distribution of proteins via gel electrophoresis, and fatty acids analysis of the lipid fraction. The antibacterial activity of all fractions was determined against Escherichia coli F4 and Streptococcus suis, two common swine pathogens. Finally, the prebiotic activity of the whole larvae and of their protein isolate and chitin-rich pellet was determined for Limosilactobacillus reuteri, a naturally occurring species in the gastro-intestinal tract of the pig. The results on amino acid profile showed that the protein extraction method used was not fully effective in cleaving chitin-bound proteins, as expected, as the true protein content of the chitin-rich sample was 48.5%. This was in accordance with the SDS-PAGE analysis, where the molecular weight distribution of proteins in the protein isolate sample did not show band at higher molecular weight, which were instead present in the chitin-rich sample. The analysis of fatty acid (FA) profile showed that lauric acid was the most abundant FA, in accordance with the literature. The results on microbiological analyses highlighted how all nitrogen-containing samples showed antibacterial activity against E. coli, the highest being the defatted fraction at 1% with a decrease of −0.77 log CFU/mL, while only the lipid fraction against S. suis (−0.48 log CFU/mL at 1%). All tested samples showed prebiotic activity for L. reuteri, but it was noticed that the increase in chitin concentration resulted in the decrease of the prebiotic effect. By combining the results of both antibacterial and prebiotic tests, it could be inferred that chitin might be considered as an antibacterial compound.

Effect of the Association of Levels of Potassium and Nitrogen from Coated Urea on the Forage Potential of Cactus Pear in Brazilian Savannah Soil

ABSTRACT The development of methods that aim to increase the water use efficiency in agricultural systems should be part of the objectives of research, and the cultivation of plants of lower water requirements, such as cactus pear, can be a good option. The purpose of this study was to evaluate the effect of the association of potassium (K) and nitrogen (N) levels from coated urea, in two harvest years, on the agronomic characteristics and nutritive value of Nopalea cochenillifera cv Doce. A randomized block design in a 2 × 4 × 2 factorial arrangement was adopted. The factors corresponded to two levels of K (40 kg and 60 kg ha−1 year−1), four levels of N (0, 60, 120, and 240 kg ha−1 year−1) and two harvest years (Years I and II). The plants were evaluated for growth, yield, chemical, and mineral composition, and nutritive value. The levels of K affected only plant height and K content in the plant. The agronomic variables and contents of ether extract and crude protein (CP) linearly increased along with the levels of N from the coated urea. There was a significant effect of interaction between N levels and harvest year on the contents of NDF, NDFap, NFC, fraction A+B1, Ca, and Mg. The level of 40 kg K ha−1 year−1 associated with 240 kg N ha−1 year−1 from coated urea provided the best results for the characteristics of growth, yield, and nutritive value of the cactus pear.

Irradiation-induced modifications of apple seed protein isolate: Exploring techno-functional, structural, thermal, and morphological characteristics

Waste valorization of apple seeds for protein extraction is a sustainable approach to the management of apple seed waste in the apple processing industry. The purpose of the research is to study the modification of apple seed protein isolates under different doses of irradiation on functional, structural, thermal, and morphological characteristics. Different irradiation doses of 8, 15, 20 25, and 50 kGy showed significant variations in protein content (88.70%–86.45%), total phenolic content (774.13–285.42 mg GAE/100g), and color profile of the apple seed protein. Irradiation altered the apple seed protein isolates' functional properties with increased foaming properties and emulsifying properties and decreased water and oil absorption properties. Irradiation induces denaturation and cross-linking in the apple seed protein structures, which increase surface hydrophobicity (708.66–1740.66), decreased zeta potential (−18.60 to −39.39 mV), and alteration in protein secondary structures with decreased α-helix (11.52–10.43 %), β-sheet content (60.82–60.13%), and random coils (16.82–16.19%), and increased β turns (10.79–13.22 %). DSC and TGA analysis revealed improved thermal properties using irradiation doses of 8–20 kGy whereas reduction for 25–50 kGy in irradiated apple seed proteins. Scanning electronic microscopy showed pores and aggregated structures of irradiated apple seed protein isolates. The study revealed the potential of irradiation as one of the non-thermal techniques for modifying apple seed protein isolates for protein valorization in various food applications.

Effects of Bergamot (Citrus bergamia Risso) By-Product on Growth Performance and Meat Quality of Growing Rabbits.

This study aimed to investigate the effects of feeding dried bergamot pulp to rabbits on animal performance and meat quality. Thirty rabbits were assigned to two groups (balanced for body weight, 804.4 ± 2.35 g) and fed individually for 60 days a basal diet (control) or the basal diet in which part of the cereals was replaced with 10% of dried bergamot pulp (DBP). There were no effects of DBP on growth performance, carcass yield, or the crude protein and ether extract composition of meat. The concentrations of α-linolenic acid (C18:3 n-3) and eicosapentaenoic acid (C20:5 n-3) increased in the longissimus thoracis et lumborum muscle (p < 0.01 and p = 0.021, respectively) after integrating dried bergamot pulp into the diet, leading to higher levels of total of ω-3 fatty acids (p < 0.01) compared to the control treatment. The inclusion of dried bergamot pulp improved the oxidative stability in meat (p < 0.001), where TBARS values were lower after 4 and 7 days of refrigerated storage (p < 0.001) in the DBP group than in the control group. Finally, feeding dried bergamot pulp to rabbits improves meat quality without negatively influencing growth performance.

Integration of MALDI-TOF MS and machine learning to classify enterococci: A comparative analysis of supervised learning algorithms for species prediction

This research focused on distinguishing distinct matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectral signatures of three Enterococcus species. We evaluated and compared the predictive performance of four supervised machine learning algorithms, K-nearest neighbor (KNN), support vector machine (SVM), and random forest (RF), to accurately classify Enterococcus species. This study involved a comprehensive dataset of 410 strains, generating 1640 individual spectra through on-plate and off-plate protein extraction methods. Although the commercial database correctly identified 76.9% of the strains, machine learning classifiers demonstrated superior performance (accuracy 0.991). In the RF model, top informative peaks played a significant role in the classification. Whole-genome sequencing showed that the most informative peaks are biomarkers connected to proteins, which are essential for understanding bacterial classification and evolution. The integration of MALDI-TOF MS and machine learning provides a rapid and accurate method for identifying Enterococcus species, improving healthcare and food safety.

Optimization of ultrasound-assisted extraction of faba bean protein isolate: Structural, functional, and thermal properties. Part 2/2

Environmental concerns linked to animal-based protein production have intensified interest in sustainable alternatives, with a focus on underutilized plant proteins. Faba beans, primarily used for animal feed, offer a high-quality protein source with promising bioactive compounds for food applications. This study explores the efficacy of ultrasound-assisted extraction under optimal conditions (123 W power, 1:15 g/mL solute/solvent ratio, 41 min sonication, 623 mL total volume) to isolate faba bean protein (U-FBPI). The ultrasound-assisted method achieved a protein extraction yield of 19.75 % and a protein content of 92.87 %, outperforming the control method’s yield of 16.41 % and protein content of 89.88 %. Electrophoretic analysis confirmed no significant changes in the primary structure of U-FBPI compared to the control. However, Fourier-transform infrared spectroscopy revealed modifications in the secondary structure due to ultrasound treatment. The U-FBPI demonstrated superior water and oil holding capacities compared to the control protein isolate, although its foaming capacity was reduced by ultrasound. Thermal analysis indicated minimal impact on the protein’s thermal properties under the applied ultrasound conditions. This research highlights the potential of ultrasound-assisted extraction for improving the functional properties of faba bean protein isolates, presenting a viable approach for advancing plant-based food production and contributing to sustainable protein consumption.

Quercetin Supplementation Improves Intestinal Digestive and Absorptive Functions and Microbiota in Rats Fed Protein-Oxidized Soybean Meal: Transcriptomics and Microbiomics Insights.

To clarify the nutritional mechanisms of quercetin mitigation in the digestive and absorptive functions in rats fed protein-oxidized soybean meal, 48 three-week-old male SD rats were randomly allocated into a 2 × 2 factorial design with two soybean meal types (fresh soybean meal or protein-oxidized soybean meal) and two quercetin levels (0 or 400 mg/kg) for a 28-day feeding trial. The protein-oxidized soybean meal treatment decreased (p < 0.05) the relative weights of the pancreas, stomach, and cecum, duodenal villus height, pancreatic and jejunal lipase activities, apparent ileal digestibility of amino acids, and apparent total tract digestibility of dry matter, crude protein, and ether extract. The supplementation of quercetin in the protein-oxidized soybean meal diet reversed (p < 0.05) the decreases in the duodenal length, ileal villus height, lipase activity, apparent ileal digestibility of amino acids, and apparent total tract digestibility of dry matter, crude protein, and ether extract. Transcriptomics revealed that the "alanine transport" and "lipid digestion and absorption" pathways were downregulated by the protein-oxidized soybean meal compared with fresh soybean meal, while the "basic amino acid transmembrane transporter activity" and "lipid digestion and absorption" pathways were upregulated by the quercetin supplementation. Microbiomics revealed that the protein-oxidized soybean meal increased the protein-degrading and inflammation-triggering bacteria in the cecum, while the relative abundances of beneficial bacteria were elevated by the quercetin supplementation.

Evaluation of Cytotoxicity and Acute Oral Toxicity of Saline Extract and Protein-Rich Fraction from Moringa oleifera Lam. Leaves.

Moringa oleifera Lam. (horseradish tree) leaves demonstrate high nutritional value, are rich in proteins, and are widely used in folk medicine and food. This study investigated the presence of secondary metabolites and antinutritional proteins in leaf extract (LE) and the protein-rich fraction (PRF) derived from M. oleifera leaves, as well as the cytotoxicity to human cells, hemolytic activity, and in vivo acute toxicity and genotoxicity in mice. The flavonoids rutin and vitexin as well as trypsin inhibitors and lectins were detected in LE and PRF. Neither sample demonstrated toxicity against human peripheral blood mononuclear cells and both showed low hemolytic action. In vivo, LE and PRF did not show antinutritional effects and caused no death. The hematological parameters of the animals in the treated group were similar to those of the control. A significant increase in the serum levels of alanine aminotransferase and a discrete leukocyte infiltration with cytoplasmic vacuolization of the hepatocytes in the liver were detected in LE-treated animals. The preparations were not genotoxic or mutagenic. This study shows that LE and PRF are not antinutritional agents and presented low acute toxicity and no genotoxicity or mutagenicity. The present study contributes to the determination of the safety of using M. oleifera leaf proteins.

Development of a Combined Protein and Dye Extraction Approach for the Analysis of Keratin-Based Textiles.

Archaeological textiles represent precious remains from ancient culture; this is because of the historical and cultural importance of the information that can be obtained by such relics. However, the extremely complicated state of preservation of these textiles, which can be charred, partially or totally mineralized, with heavy soil or biological contamination, requires highly specialized and sensitive analytical tools to perform a comprehensive study. Starting from these considerations, the paper presents a combined workflow that provides the extraction of dyes and keratins and keratin-associated proteins in a single step, minimizing sampling while maximizing the amount of information gained. In the first phase, different approaches were tested and two different protocols were found suitable for the purpose of the unique workflow for dyes/keratin-proteins: a slightly modified urea protocol and a recently proposed new TCEP/CAA procedure. In the second step, after the extraction, different methods of cleanup and workflow for proteins and dyes were investigated to develop protocols that did not result in a loss of aliquots of the analytes of interest and to maximize the recovery of both components from the extracting solution. These protocols investigated the application of two types of paramagnetic beads, unmodified and carboxylate-coated hydrophilic magnetic beads, and dialysis and stage-tip protocols. The newly designed protocols have been applied to cochineal, weld, orchil, kermes, and indigo keratin-based dyed samples to evaluate the effectiveness of the protocols on several dye sources. These protocols, based on a single extraction step, show the possibility of investigating dyes and keratins from a unique sample of 1 mg or lesser, with respect to the thresholds of sensitivity and accuracy required in the study of textile artifacts of historical and artistic values.

Leech Poecilobdella manillensis protein extract ameliorated hyperuricemia by restoring gut microbiota dysregulation and affecting serum metabolites.

Hyperuricemia (HUA) is a public health concern that needs to be solved urgently. The lyophilized powder of Poecilobdella manillensis has been shown to significantly alleviate HUA; however, its underlying metabolic regulation remains unclear. To explore the underlying mechanisms of Poecilobdella manillensis in HUA based on modulation of the gut microbiota and host metabolism. A mouse model of rapid HUA was established using a high-purine diet and potassium oxonate injections. The mice received oral drugs or saline. Additionally, 16S rRNA sequencing and ultra-high performance liquid chromatography with quadrupole time-of-flight mass spectrometry-based untargeted metabolomics were performed to identify changes in the microbiome and host metabolome, respectively. The levels of uric acid transporters and epithelial tight junction proteins in the renal and intestinal tissues were analyzed using an enzyme-linked immunosorbent assay. The protein extract of Poecilobdella manillensis lyophilized powder (49 mg/kg) showed an enhanced anti-trioxypurine ability than that of allopurinol (5 mg/kg) (P < 0.05). A total of nine bacterial genera were identified to be closely related to the anti-trioxypurine activity of Poecilobdella manillensis powder, which included the genera of Prevotella, Delftia, Dialister, Akkermansia, Lactococcus, Escherichia_Shigella, Enterococcus, and Bacteroides. Furthermore, 22 metabolites in the serum were found to be closely related to the anti-trioxypurine activity of Poecilobdella manillensis powder, which correlated to the Kyoto Encyclopedia of Genes and Genomes pathways of cysteine and methionine metabolism, sphingolipid metabolism, galactose metabolism, and phenylalanine, tyrosine, and tryptophan biosynthesis. Correlation analysis found that changes in the gut microbiota were significantly related to these metabolites. The proteins in Poecilobdella manillensis powder were effective for HUA. Mechanistically, they are associated with improvements in gut microbiota dysbiosis and the regulation of sphingolipid and galactose metabolism.

Comparative Proteomic Study of Retinal Ganglion Cells Undergoing Various Types of Cellular Stressors.

Retinal ganglion cell (RGC) damage serves as a key indicator of various retinal degenerative diseases, including diabetic retinopathy (DR), glaucoma, retinal arterial and retinal vein occlusions, as well as inflammatory and traumatic optic neuropathies. Despite the growing body of data on the RGC proteomics associated with these conditions, there has been no dedicated study conducted to compare the molecular signaling pathways involved in the mechanism of neuronal cell death. Therefore, we launched the study using two different insults leading to RGC death: glutamate excitotoxicity and optic nerve crush (ONC). C57BL/6 mice were used for the study and underwent NMDA- and ONC-induced damages. Twenty-four hours after ONC and 1 hour after NMDA injection, we collected RGCs using CD90.2 coupled magnetic beads, prepared protein extracts, and employed LC-MS for the global proteomic analysis of RGCs. Statistically significant changes in proteins were analyzed using the Shiny Go program to identify GO biological processes and molecular functions resulting from the treatment. We identified unique and common alterations in protein profiles in RGCs undergoing different types of cellular stressors. Additionally, we observed the absence of certain proteins in treated RGCs compared to the control group. Our study not only identified both unique and shared proteomic changes but also laid the groundwork for the future development of a therapeutic platform for testing gene candidates for DR and glaucoma.

A proteome-wide quantitative platform for nanoscale spatially resolved extraction of membrane proteins into native nanodiscs.

The intricate molecular environment of the native membrane profoundly influences every aspect of membrane protein (MP) biology. Despite this, the most prevalent method of studying MPs uses detergent-like molecules that disrupt and remove this vital local membrane context. This severely impedes our ability to quantitatively decipher the local molecular context and comprehend its regulatory role in the structure, function, and biogenesis of MPs. Using a library of membrane-active polymers we have developed a platform for the high-throughput analysis of the membrane proteome. The platform enables near-complete spatially resolved extraction of target MPs directly from their endogenous membranes into native nanodiscs that maintain the local membrane context. We accompany this advancement with an open-access database that quantifies the polymer-specific extraction variability for 2065 unique mammalian MPs and provides the most optimized condition for each of them. Our method enables rapid and near-complete extraction and purification of target MPs directly from their endogenous organellar membranes at physiological expression levels while maintaining the nanoscale local membrane environment. Going beyond the plasma membrane proteome, our platform enables extraction from any target organellar membrane including the endoplasmic reticulum, mitochondria, lysosome, Golgi, and even transient organelles such as the autophagosome. To further validate this platform, we took several independent MPs and demonstrated how our resource can enable rapid extraction and purification of target MPs from different organellar membranes with high efficiency and purity. Further, taking two synaptic vesicle MPs, we show how the database can be extended to capture multiprotein complexes between overexpressed MPs. We expect these publicly available resources to empower researchers across disciplines to efficiently capture membrane 'nano-scoops' containing a target MP and interface with structural, functional, and other bioanalytical approaches. We demonstrate an example of this by combining our extraction platform with single-molecule TIRF imaging to demonstrate how it can enable rapid determination of homo-oligomeric states of target MPs in native cell membranes.

Response surface methodology guided approach for optimization of protein isolate from Faba bean. Part 1/2

Ultrasound-assisted extraction (UAE) was evaluated as a green procedure to produce faba beans protein isolates from faba beans. Magnetic stirring was performed as conventional extraction. A three-level five-factor Box-Behnken Design (BBD) was applied to obtain the optimal UAE conditions to concurrently maximize extraction yield and protein content. The response surface methodology (RSM) showed a quadratic curvature for extraction yield and protein. The optimal extraction conditions were determined as: Power of 123 W, solute/solvent ratio of 0.06 (1:15 g/mL), sonication time of 41 min, and total volume of 623 mL with a desirability value of 0.82. Under these conditions, the extraction yield of 19. 75 ± 0.87 % (Protein yield of 67.84 %) and protein content of 92.87 ± 0.53 % were obtained for optimum ultrasound extraction. Control samples using magnetic stirring under similar conditions without ultrasound treatment showed an extraction yield of 16.41 ± 0.02 % (Protein yield of 54.65 %) and a protein content of 89. 88 ± 0.40 %. This shows that BBD can effectively be used to optimize the extraction of proteins from faba beans using optimal extraction conditions, resulting in a higher extraction yield and protein purity.

RETRACTION: Nuclear Clusterin Accumulation during Heat Shock Response: Implications for Cell Survival and Thermo-tolerance Induction in Immortalized and Prostate Cancer Cells.

A. E. Caccamo, S. Desenzani, L. Belloni, A. F. Borghetti, S. Bettuzzi, "Nuclear Clusterin Accumulation during Heat Shock Response: Implications for Cell Survival and Thermo-tolerance Induction in Immortalized and Prostate Cancer Cells," Journal of Cellular Physiology 207, no. 1 (2006): 208-219, https://doi.org/10.1002/jcp.20561. The above article, published online on 5 December 2005 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the journal Editor-in-Chief, Alexander Hutchison; and Wiley Periodicals LLC. The retraction has been agreed following allegations raised by third parties. Several lanes within the Western Blot images displayed in Figures 1 A, 2 A, 2B, and 7 were found to display highly similar patterns despite depicting protein extracts from samples of different origin/treatment. As the raw data could not be retrieved due to the significant time elapsed since publication, the concerns could not be addressed. In light of the significant number of concerns, the editors have lost trust in the overall accuracy of the data presented. The corresponding author S. Bettuzzi disagrees with the decision of retraction. Confirmation of retraction could not be obtained by the remaining co-authors.