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Plants, as sessile organisms, must adapt to a range of abiotic stresses, including drought, salinity, heat, and cold, which are increasingly exacerbated by climate change. These stresses significantly impact crop productivity, posing challenges for sustainable agriculture and food security. Recent advances in omics studies and genetics have shed light on molecular mechanisms underlying plant stress responses, including the role of calcium (Ca2⁺) signaling, liquid–liquid phase separation (LLPS), and cell wall-associated sensors in detecting and responding to environmental changes. However, gaps remain in understanding how rapid stress signaling is integrated with slower, adaptive processes. Emerging evidence also highlights crosstalk between abiotic stress responses, plant immunity, and growth regulation, mediated by key components such as RAF-SnRK2 kinase cascades, DELLA proteins, etc. Strategies to enhance crop stress resistance without compromising yield include introducing beneficial alleles, spatiotemporal optimization of stress responses, and decoupling stress signaling from growth inhibition. This review emphasizes the importance of interdisciplinary approaches and innovative technologies to bridge fundamental research and practical agricultural applications, aiming to develop resilient crops for sustainable food production in an era of escalating environmental challenges. [ABSTRACT FROM AUTHOR]

Oxidative stress is widely recognized as a key mechanism in the development of hypertension. Under pathological conditions, such as in hypertension, oxidative stress leads to irreversible posttranslational modifications of proteins, which result in loss of protein function and cellular damage. We have previously documented physiological and morphological changes across various blood and bone marrow cell lineages, all of which exhibit elevated oxidative stress. While cytoskeletal changes in erythrocytes have been well characterized in hereditary diseases, this is the first study, to our knowledge, to investigate cytoskeletal reorganization in erythrocytes from hypertensive patients. To this end, we compared the expression patterns and subcellular distribution of key cytoskeletal proteins in erythrocytes from hypertensive individuals with those from normotensive subjects using Western blot, flow cytometry, and confocal microscopy. Our results revealed the presence of three erythrocyte subpopulations with differential expression of glycophorin A. The persistent oxidative environment in hypertensive patients causes dysregulation in the expression of glycophorin A, Band 3 protein, protein 4.1, and ankyrin, as well as the reorganization of spectrin. These alterations in protein expression and distribution suggest that oxidative stress in hypertensive individuals may induce structural modifications, ultimately impairing erythrocyte membrane elasticity and function. [ABSTRACT FROM AUTHOR]

Auxenochlorella pyrenoidosa is a microalga that stands out due to its high protein content. The objective of this work was to study the effect of high-pressure homogenization (HPH) on the recovery of proteins from A. pyrenoidosa and their application as functional emulsifiers. Untreated and HPH-treated (400–800 bar, 1 and 4 passes) aqueous cellular suspensions were incubated at 40 °C for 6 h. The aqueous extracts were collected, the proteins were precipitated at pH 3, and the Auxenochlorella pyrenoidosa protein concentrates (APPC) were lyophilized. Increasing HPH pressure and number of passes (400–800 bar, 1 and 4 passes) improved protein recovery yield up to 57%. Higher HPH pressures also reduced α-helix and β-sheet structures, exposing the hydrophobic protein core. This protein modification led to APPCs with increased oil-holding capacity (2.83 g oil/g APPC). The surface tension of APPC solutions reached a minimum value of 28.6 mN/m at an APPC concentration of 2% w/w. The APPCs from untreated and HPH-treated biomass were used to stabilize nanoemulsions (2–6% sunflower oil), comparing one-step homogenization (high-speed homogenization) with the two-step homogenization method (combining high-speed and high-pressure homogenization). The two-step method led to significantly smaller oil droplets with narrow size distribution, leading to stable nanoemulsions with improved resistance to centrifugation and heating–cooling cycles. Due to APPC's great emulsifying properties, A. pyrenoidosa proteins have a promising potential for various applications such as delivery systems stabilization. Additionally, the low energy requirements, continuous processing capability, and scalability of HPH make it a suitable process for industrial applications. [ABSTRACT FROM AUTHOR]

Glycopeptides derived from the glycoprotein mucin‐1 (MUC1) have shown potential as tumor‐associated antigens for cancer vaccine development. However, their low immunogenicity and non‐selective conjugation to carriers present significant challenges for the clinical efficacy of MUC1‐based vaccines. Here, we introduce a novel vaccine candidate based on a structure‐guided design of an artificial antigen derived from MUC1 glycopeptide. This engineered antigen contains two non‐natural amino acids and has an α‐S‐glycosidic bond, where sulfur replaces the conventional oxygen atom linking the peptide backbone to the sugar N‐acetylgalactosamine. The glycopeptide is then specifically conjugated to the immunogenic protein carrier CRM197 (Cross‐Reactive Material 197), a protein approved for human use. Conjugation involves selective reduction and re‐bridging of a disulfide in CRM197, allowing the attachment of a single copy of MUC1. This strategy results in a chemically defined vaccine while maintaining both the structural integrity and immunogenicity of the protein carrier. The vaccine elicits a robust Th1‐like immune response in mice and generates antibodies capable of recognizing human cancer cells expressing tumor‐associated MUC1. When tested in mouse models of colon adenocarcinoma and pancreatic cancer, the vaccine is effective both as a prophylactic and therapeutic use, significantly delaying tumor growth. In therapeutic applications, improved outcomes were observed when the vaccine was combined with an anti‐programmed cell death protein 1 (anti‐PD‐1) checkpoint inhibitor. Our strategy reduces batch‐to‐batch variability and enhances both immunogenicity and therapeutic potential. This site‐specific approach disputes a prevailing dogma where glycoconjugate vaccines require multivalent display of antigens. [ABSTRACT FROM AUTHOR]

The functionalization of protein sidechains with highly water-soluble chlorotriazines (or derivatives thereof) using nucleophilic aromatic substitution reactions has been commonly employed to install various functional groups, including poly(ethylene glycol) tags or fluorogenic labels. Here, a poorly soluble dichlorotriazine with an appended indole is shown to react with a construct containing the disordered domain of BRCA1. Subsequently, this construct can undergo proteolytic cleavage to remove the SUMO-tag: the N-terminal poly(His) tag is still effective for purification. Steady-state fluorescence, circular dichroism spectroscopy, and isothermal titration calorimetry with the binding partner of BRCA1, PALB2, are used to characterize the indole-labeled BRCA1. Neither the reaction conditions nor the indole-tag appreciably alter the structure of the BRCA1. Mass spectrometry confirms that the target is modified once, although the location of modification cannot be determined by tandem mass spectrometry with collision-induced dissociation due to disadvantageous fragmentation patterns. [ABSTRACT FROM AUTHOR]

Covalent modification of proteins at specific, predetermined sites is essential for advancing biological and biopharmaceutical applications. Site-selective labeling techniques for protein modification allow us to effectively track biological function, intracellular dynamics, and localization. Despite numerous reports on modifying target proteins with functional chemical probes, unique organic reactions that achieve site-selective integration without compromising native functional properties remain a significant challenge. In this review, we delve into site-selective protein modification using synthetic probes, highlighting both chemical and computational methodologies for chemo- and regioselective modifications of naturally occurring amino acids, as well as proximity-driven protein-selective chemical modifications. We also underline recent traceless affinity labeling strategies that involve exchange/cleavage reactions and catalyst tethering modifications. The rapid development of computational infrastructure and methods has made the bioconjugation of proteins more accessible, enabling precise predictions of structural changes due to protein modifications. Hence, we discuss bioconjugational computational approaches, including molecular dynamics and artificial intelligence, underscoring their potential applications in enhancing our understanding of cellular biology and addressing current challenges in the field. [ABSTRACT FROM AUTHOR]

The native sunflower meal protein (SFMP) offers promising applications in food structuring and functionality. To enhance the feasibility and efficiency of SFMP utilization, this study investigated the effects of alkaline pH-shifting modification on its functional, physicochemical, and structural properties. The structural changes induced by alkaline pH-shifting were evaluated using Fourier-transform infrared (FTIR), X-ray diffraction (XRD), and scanning electron microscopy (SEM) analyses, revealing a significant increase in the random coil content from 5.13% in native protein (NP) to 31.38% in modified protein (MP). Furthermore, pH-shifting resulted in an expansion of the crystallization area in the 20° region and a reduction in the 10° zone. Additionally, Pickering emulsions (CNPE and CMPE) produced using chitosan were found to have higher emulsion stability compared to conventional emulsions (NPE and MPE). The samples with the highest absolute ζ-potential values were CMP (52.03 ± 0.78) and CNP (52.26 ± 1.89). Notably, CMP exhibited the best emulsion stability, with particle sizes of 19.98 ± 0.92 µm and 22.29 ± 1.03 µm on the first and fourteenth day, respectively. Overall, this study successfully demonstrated the creation of stable emulsions from SFMP isolate. [ABSTRACT FROM AUTHOR]

Powdered milk is a food ingredient valued for its long-term shelf life and space-saving qualities. However, the process of concentrating and drying milk, usually through spray drying, can lead to chemical modifications in proteins. This study aims to evaluate how two drying technologies, spray drying (SD) and pulse spray drying (PSD), affect protein modifications in skim milk powders (SMP). Batches of SMP were produced using both SD and PSD technologies, with drying outlet temperatures set at 70 °C and 100 °C. Electrophoresis, chromatography, and mass spectrometry were exploited to assess protein modifications induced by the drying processes. In vitro digestion models were applied to evaluate the digestibility scores of the SMP. The treatment temperature was the major factor for protein modifications in both PSD and SD processes, as evidenced by electrophoresis and chromatography analysis. As indicated by chromatography, PSD technology resulted in lower whey protein modifications compared to SD. Interestingly, the digestibility scores were higher than 95 and no significant differences were observed between the two technologies. In conclusion, proteomic methods effectively identified and semi-quantified thermal-induced modifications in SMP produced using an alternative drying technology as compared to the conventional SD. PSD is a convenient powdering process that preserves protein quality and functionality. [ABSTRACT FROM AUTHOR]

Low-temperature plasma (LTP) jets are Food and Drug Administration (FDA)-approved medical devices to remove cancerous tissue and aid in wound healing. However, reports on their reaction with proteins are conflicting, ranging from fragmentation, oxidation, aggregation, or a combination thereof. In this study we bridge the gap between plasma-treatment of short peptides to proteins at physiologically relevant concentrations. The LTP in this study is based on a helium dielectric barrier discharge that forms a plasma-jet, which is directed at the solution without direct contact with the plasma, and results in the formation of reactive oxygen species (ROS) OH• and O2•− in solution. The longer the solution is treated, the more solution-phase ROS form. Treating peptide- and protein-containing solutions leads to extensive oxidation. The ROS led to the same oxidative modifications for peptide M with increasing chain length (9, 18, 37, 76 amino acids), which could be identified with high-resolution mass spectrometry. Oxidized species M + x O led to conformational changes such as compaction and elongation, while the unmodified peptide M remained unaltered, as found by ion mobility spectrometry and size exclusion chromatography. For proteins at high concentration, insoluble aggregates formed and could be identified by UV/V is light scattering and atomic force microscopy. The formation of aggregates is dependent on the amino acid chain length, the peptide concentration, and the time for aggregate formation. These findings highlight the importance of both peptide chain length and concentration in determining the fate of peptides following the exposure to LTP, while also offering valuable insights for the field of plasma medicine. [ABSTRACT FROM AUTHOR]

Maleimide derivatives are privileged reagents for chemically modifying proteins through the Michael addition reaction with cysteine due to their selectivity, operational simplicity, and commercial availability. However, since accessible free cysteine is rarely found in natural proteins, it is highly desirable to find alternative targets to enable direct bioconjugation of proteins with maleimides. In this study, we have developed an operationally simple and straightforward method for the N‐terminal modification of proteins without the need for mutagenesis via a copper(II)‐mediated [3+2] cycloaddition reaction with maleimides and 2‐pyridinecarboxaldehyde (2‐PCA) derivatives under non‐denaturing conditions at pH 6 and 37 °C in aqueous media. Our method utilizes commercially available maleimides to attach diverse functionalities to various N‐terminal amino acids. We demonstrate the preparation of a ternary protein complex cross‐linked at the N‐termini and dually modified trastuzumab equipped with monomethyl auristatin E (MMAE), a cytotoxic agent, and a Cy5 fluorophore (MMAE‐Cy5‐trastuzumab). MMAE‐Cy5‐trastuzumab retained human epidermal growth factor receptor 2 (HER2) recognition activity and exerted cytotoxicity against HER2‐positive cells. Furthermore, MMAE‐Cy5‐trastuzumab allowed successful visualization of HER2‐positive cancer cells in mouse tumors. This straightforward method will expand the accessibility of protein conjugates with well‐defined structures in a wide range of research fields. [ABSTRACT FROM AUTHOR]

The functional properties of pumpkin seed proteins remain unutilized in numerous food and industrial applications. Several current approaches aim to improve the functional properties of pumpkin seed proteins, allowing their innovative potential to develop and modify significantly. Several strategies can be implemented to alter the functional properties of proteins isolated from pumpkin seeds. The first is enzymatic hydrolysis, regardless of whether, proteases may free peptide binding and profoundly impact the protein structure and functionality. Thermal treatment can include heating and cooling to replace protein conformation and increase solubility, emulsification, and gelation properties. Chemical modification techniques, including acylation and glycation, can also be used to improve stability, viscosity, and foaming ability. Functional properties and, where possible, ingredients with many applications may include exceptional possibilities for proteins modified in food preparations, such as dairy replacements, plant-based meat analogues, and free gluten that have an outstanding aspect, satisfactory quality, and nutritional profiles. As multiple different proteins act as precursors of active peptides, they can also be used to generate bio-specific foods. This review briefly provides information about various types of protein extraction techniques and functional properties that are modified by different types of processing technologies. [ABSTRACT FROM AUTHOR]

Organic halogen compounds are cornerstones of applied chemical sciences. Halogen substitution is a smart molecular design strategy adopted to influence reactivity, membrane permeability and receptor interaction. Chiral bioreceptors may restrict the stereochemical requirements in the halo‐ligand design. Straightforward (but expensive) catalyzed stereospecific halogenation has been reported. Historically, PCl5 served access to uncatalyzed stereoselective chlorination although the stereochemical outcomes were influenced by steric parameters. Nonetheless, stereochemical investigation of PCl5 reaction mechanism with carbamoyl (RCONHX) compounds has never been addressed. Herein, we provide the first comprehensive stereochemical mechanistic explanation outlining halogenation of carbamoyl compounds with PCl5; the key regioselectivity‐limiting nitrilimine intermediate (8‐Z.HCl); how substitution pattern influences regioselectivity; why oxadiazole byproduct (P1) is encountered; stereo‐electronic factors influencing the hydrazonoyl chloride (P2) production; and discovery of two stereoselectivity‐limiting parallel mechanisms (stepwise and concerted) of elimination of HCl and POCl3. DFT calculations, synthetic methodology optimization, X‐ray evidence and experimental reaction kinetics study evidence all supported the suggested mechanism proposal (Scheme 2). Finally, we provide mechanism‐inspired future recommendations for directing the reaction stereoselectivity toward elusive and stereochemically inaccessible (E)‐bis‐hydrazonoyl chlorides along with potentially pivotal applications of both (E/Z)‐stereoisomers especially in medicinal chemistry and protein modification. [ABSTRACT FROM AUTHOR]

Streptavidin is a tetrameric protein with high specificity and affinity for biotin. The interaction between avidin and biotin has become a valuable tool in nanotechnology. In recent years, the site‐specific biotin modification of proteins using biotin ligases, such as BirA, has attracted attention. This study established an in vivo method for achieving the complete biotinylation of target proteins using a single plasmid co‐expressing BirA and its target proteins. Specifically, a biotin‐modified protein was produced in Escherichia coli strain BL21(DE3) using a single plasmid containing genes encoding both BirA and a protein fused to BirA's substrate sequence, Avitag. This approach simplifies the production of biotinylated proteins in E. coli and allows the creation of various biotinylated protein types through gene replacement. Furthermore, the biotin modification rate of the obtained target protein could be evaluated using Native‐PAGE without performing complicated isolation operations of biotinylated proteins. In Native‐PAGE, biotin‐modified proteins and unmodified proteins were confirmed as clearly different bands, and it was possible to easily derive the modification rate from the respective band intensities. [ABSTRACT FROM AUTHOR]

This study investigates the impact of aqueous ozone (AO) on the yield, molecular structure, and rheological properties of wheat gluten separated using the batter procedure. Employing strong gluten flour (SGF) and weak gluten flour (WGF), we demonstrate that AO pretreatment significantly enhances the yield and purity of separated starch and gluten. Surface hydrophobicity, free sulfhydryl groups, Fourier transform infrared spectroscopy (FTIR), Raman, and size exclusion‐high‐performance liquid chromatography (SE‐HPLC) analyses were used to evaluate the effects of AO on the molecular structure of gluten. Our analysis reveals that low concentrations of AO induce specific modifications in gluten proteins. AO treatment increases cross‐linking in glutenin macropolymer (GMP), reduces surface hydrophobicity, and stabilizes secondary and tertiary structures. These changes include an increase in β‐sheet content by approximately 9% and a corresponding decrease in β‐turn structures, leading to enhanced viscoelastic properties of the gluten. The research highlights AO's potential as a sustainable and efficient agent in wheat flour processing, offering advancements in both product quality and eco‐friendly processing techniques. Future research should optimize AO treatment parameters and explore its effects on different cereal types further to enhance its applicability and benefits in food processing. Practical Application: Our work substantially advances the existing knowledge on wheat flour processing by demonstrating the multifaceted benefits of AO pretreatment. We unveil significant improvements in the yield and purity of starch and gluten when compared to conventional separation methods. Moreover, our in‐depth analysis of molecular changes induced by AO, including increased cross‐linking, alterations in surface hydrophobicity, and modifications in glutenin macropolymer content, provides new insights into how AO affects the viscoelastic properties of gluten. This contribution is pivotal for the development of more efficient, sustainable, and eco‐friendly wheat flour processing technologies. [ABSTRACT FROM AUTHOR]

Summary: In this study, ultrasound‐assisted covalent binding method, free radical‐induced method, was used to prepare Moringa protein isolate (MPI) – gallic acid conjugates (MPI‐GAC) to examine the improvement of the sonication application. Additionally, the influence of polyphenol conjugation on functional and structural attributes of MPI was assessed thru spectral analysis, scanning electron microscopy, antioxidant activity, emulsification, foaming, and water and oil holding properties. The enhanced value of grafted polyphenol species was observed after introducing ultrasound treatment during free radical mediation process. The structural changes in protein by gallic acid conjugation were confirmed by using FT‐IR which illustrated the modification in MPI‐GAC with modified peak width of the amide‐I band. Scanning electron microscopy also showed a significant difference in the structure of MPI and MPI‐GAC. The grafting of gallic acid onto MPI directed under ultrasound‐assisted conditions conferred distinctive functional characteristics, containing elevated emulsification capacity (51.4 mg mL−1) along with enhanced water and oil holding capacity by 2.07 and 5.11 g g−1, respectively. Furthermore, conjugation also increased the protein solubility 89.3–97.6% and significant trend (68.04–88.06%) of antioxidant capacity was observed by increasing the concentration. The findings of this research work offer new insights on the effective fabrication of MPI‐GAC with multifaceted functionalities, thus broadening the scope and possibilities of high‐value Moringa seed protein. [ABSTRACT FROM AUTHOR]

Key message: Analysis of the N-terminome of Physcomitrella reveals N-terminal monomethylation of nuclear-encoded, mitochondria-localized proteins. Post- or co-translational N-terminal modifications of proteins influence their half-life as well as mediating protein sorting to organelles via cleavable N-terminal sequences that are recognized by the respective translocation machinery. Here, we provide an overview on the current modification state of the N-termini of over 4500 proteins from the model moss Physcomitrella (Physcomitrium patens) using a compilation of 24 N-terminomics datasets. Our data reveal distinct proteoforms and modification states and confirm predicted targeting peptide cleavage sites of 1,144 proteins localized to plastids and the thylakoid lumen, to mitochondria, and to the secretory pathway. In addition, we uncover extended N-terminal methylation of mitochondrial proteins. Moreover, we identified PpNTM1 (P. patens alpha N-terminal protein methyltransferase 1) as a candidate for protein methylation in plastids, mitochondria, and the cytosol. These data can now be used to optimize computational targeting predictors, for customized protein fusions and their targeted localization in biotechnology, and offer novel insights into potential dual targeting of proteins. [ABSTRACT FROM AUTHOR]

Polysulfides are significant compounds in life science, pharmaceutical science, and materials science. Therefore, polysulfide construction is in great demand. The controllable sequential installation of groups on both ends of a S−S motif faces an enormous challenge owing to the reversible nature of the covalent S−S bond. A library was established with two divergent mask groups for bilateral unsymmetrical disulfurating reagents (R1O−SS−SO2R2). Sequential coupling with preferential activation of the S−SO2 bond (37.6 kcal/mol) and controllable activation of the S−O bond (54.8 kcal/mol) in the presence of the S−S bond (62.0 kcal/mol) enabled successive reactions at each end of the S−S motif to afford unsymmetrical disulfides and trisulfides, even for the cross‐linkage of natural products, pharmaceuticals, peptides, and a protein (bovine serum albumin). [ABSTRACT FROM AUTHOR]

Chemical modifications of proteins induced by ambient ozone (O 3) and nitrogen oxides (NO x) are of public health concerns due to their potential to trigger respiratory diseases. The laboratory and environmental exposure systems have been widely used to investigate their relevant mechanism in the atmosphere. Using bovine serum albumin (BSA) as a model protein, we evaluated the two systems and aimed to reduce the uncertainties of both the reactants and products in the corresponding kinetic study. In the laboratory simulation system, the generated gaseous pollutants showed negligible losses. Ten layers of BSA were coated on the flow tube with protein extraction recovery of 87.4%. For environmental exposure experiment, quartz fiber filter was selected as the upper filter with low gaseous O 3 (8.0%) and NO 2 (1.7%) losses, and cellulose acetate filter was appropriate for the lower filter with protein extraction efficiency of 95.2%. The protein degradation process was observed without the exposure to atmospheric oxidants and contributed to the loss of protein monomer mass fractions, while environmental factors (e.g., molecular oxygen and ultraviolet) may cause greater protein monomer losses. Based on the evaluation, the study exemplarily applied the two systems to protein modification and both showed that O 3 promotes the protein oligomerization and nitration, while increased temperature can accelerate the oligomerization and increased relative humidity can inhibit the nitration in the environmental exposure samples. The developed laboratory and environmental systems are suitable for studying protein modifications formed under different atmospheric conditions. A combination of the two will further reveal the actual mechanism of protein modifications. [Display omitted] [ABSTRACT FROM AUTHOR]

为促进花生蛋白的深加工和更广泛的应用，采用高静压联合碱性蛋白酶酶法改性花生蛋白。通过单因素试验考察了静压力、pH、酶添加量、酶解时间和酶解温度对花生蛋白溶解度的影响，在此基础上，采用正交试验优化花生蛋白联合改性工艺条件，并测定了联合改性花生蛋白的起泡性和泡沫稳定性、巯基和二硫键含量以及总还原能力。结果表明：花生蛋白联合改性最佳工艺条件为静压力300 MPa、1 g/100 mL碱性蛋白酶（20万U/g）添加量3.0 mL（100 mL质量分数5%的花生蛋白溶液）、酶解时间60 min、pH 10、酶解温度50 ℃，在此条件下联合改性花生蛋白溶解度为（82.87±0.51）%；联合改性花生蛋白的起泡性、泡沫稳定性、巯基含量、总还原能力显著提高，二硫键含量显著下降。综上，高静压联合酶法改性改善了花生蛋白的理化性质及功能特性，有利于其深加工及更广泛的应用。In order to promote the deep processing and broader application of peanut protein, peanut protein was modified by high hydrostatic pressure combined with alkaline protease hydrolysis. The effects of hydrostatic pressure, pH, enzyme addition amount, enzymatic time and enzymatic temperature on the solubility of peanut protein were investigated by single factor experiment, then orthogonal experiment was used to optimize the modification process of peanut protein. The foaming property and foam stability, sulfhydryl and disulfide bond contents, and total reducing capacity of the co-modified peanut protein were determined. The results showed that the optimal co-modification conditions for peanut protein were hydrostatic pressure 300 MPa, 1 g/100 mL alkaline protease(200 000 U/g) addition amount 3.0 mL (based on 100 mL peanut protein solution with mass fraction of 5%), pH 10, enzymatic temperature 50 ℃, and enzymatic time 60 min, and the protein solubility was (82.87±0.51)% under these conditions. The foaming property and foam stability, sulfhydryl group content, and total reducing capacity of co-modified peanut protein improved, while the disulfide bond content decreased. In conclusion, the combination of high hydrostatic pressure and enzymatic method can improve the physicochemical properties and functional properties of peanut protein, which facilitates the deep processing and broader application of peanut protein.

Nitric oxide (NO) induces protein posttranslational modification (PTM), known as S-nitrosylation, which has started to gain attention as a critical regulator of thousands of substrate proteins. However, our understanding of the biological consequences of this emerging PTM is incomplete because of the limited number of identified S-nitrosylated proteins (S–NO proteins). Recent advances in detection methods have effectively contributed to broadening the spectrum of discovered S–NO proteins. This article briefly reviews the progress in S–NO protein detection methods and discusses how these methods are involved in characterizing the biological consequences of this PTM. Additionally, we provide insight into S–NO protein-related diseases, focusing on the role of these proteins in mitigating the severity of infectious diseases. [ABSTRACT FROM AUTHOR]

Lysine‐specific peptide and protein modification strategies are widely used to study charge‐related functions and applications. However, these strategies often result in the loss of the positive charge on lysine, significantly impacting the charge‐related properties of proteins. Herein, we report a strategy to preserve the positive charge and selectively convert amines in lysine side chains to amidines using nitriles and hydroxylamine under aqueous conditions. Various unprotected peptides and proteins were successfully modified with a high conversion rate. Moreover, the reactive amidine moiety and derived modification site enable subsequent secondary modifications. Notably, positive charges were retained during the modification. Therefore, positive charge‐related protein properties, such as liquid‐liquid phase separation behaviour of α‐synuclein, were not affected. This strategy was subsequently applied to a lysine rich protein to develop an amidine‐containing coacervate DNA complex with outstanding mechanical properties. Overall, our innovative strategy provides a new avenue to explore the characteristics of positively charged proteins. [ABSTRACT FROM AUTHOR]

Summary: Proteins are commonly used ingredients to impart certain non‐nutritional properties to foods. Consequently, the industry is constantly seeking to enhance proteins that provide these attributes to food products. Therefore, the aim of this research was to extract the protein from sesame seeds and evaluate the effect of chemical modification through acylation with lauroyl chloride and acetic anhydride on functional properties (solubility index, emulsifying, foaming, swelling and water‐holding and oil‐holding capacity). Chemical modification was successfully demonstrated using FTIR spectroscopy and X‐rays. It was found that acylation with lauroyl chloride or acetic anhydride altered the physicochemical and functional properties, particularly at pH 10. Both reactions exhibited substitution degrees exceeding 20%. Chemical modification via acylation resulted in significant improvements in water and oil retention capacity, emulsifying activity and foaming activity. Thermal analysis revealed significant changes in the modified samples, altering their denaturation temperature, leading to the conclusion that chemical modifications through acylation with lauroyl chloride or acetic anhydride offer benefits in certain functional properties relevant to the food industry. They also represent an opportunity to diversify the application of a by‐product obtained from sesame oil extraction. [ABSTRACT FROM AUTHOR]

The present study demonstrates the effects of pH-shifting treatments and magnetic field-assisted pH-shifting treatments on the properties of myofibrillar protein (MP) in frozen meat. The solubility results indicate that the pH-shifting treatments increased the solubility of MP from 16.8% to a maximum of 21.0% (pH 9). The values of surface hydrophobicity and protein particle size distribution indicate that the pH-shifting treatment effectively inhibited protein aggregation through electrostatic interactions. However, under higher pH conditions (pH 10, 11), the treatments assisted by the magnetic field increased the degree of aggregation. The total thiol content and SDS-PAGE results further suggest that the magnetic field-assisted pH-shifting treatment accelerated the formation of covalent bonds among MPs under the alkaline environment. The results of the Differential Scanning Calorimetry (DSC) and protein secondary structure analysis indicate that the magnetic field promoted the unfolding of protein structures in an alkaline environment, markedly reducing the effective pH levels of pH-shifting. Electron paramagnetic resonance (EPR) data indicate that the phenomenon might be associated with the increased concentration of free radicals caused by the magnetic field treatment. In summary, the application of magnetic field-assisted pH-shifting treatments could emerge as a potent and promising strategy to improve the protein properties in frozen meat. [ABSTRACT FROM AUTHOR]

This review presents a comprehensive analysis of plant-based proteins from soybeans, pulses, cereals, and pseudo-cereals by examining their structural properties, modification techniques, bioactivities, and applicability in food systems. It addresses the critical need for a proper utilization strategy of proteins from various plant sources amidst the rising environmental footprint of animal protein production. The inherent composition diversity among plant proteins, their nutritional profiles, digestibility, environmental impacts, and consumer acceptance are compared. The innovative modification techniques to enhance the functional properties of plant proteins are also discussed. The review also investigates the bioactive properties of plant proteins, including their antioxidant, antimicrobial, and antitumoral activities, and their role in developing meat analogs, dairy alternatives, baked goods, and 3D-printed foods. It underscores the consideration parameters of using plant proteins as sustainable, nutritious, and functional ingredients and advocates for research to overcome sensory and functional challenges for improved consumer acceptance and marketability. [ABSTRACT FROM AUTHOR]

Secretoglobin (SCGB) 3A2 belongs to an intriguing family of small, secreted proteins present only in mammals. Although members of the SCGB protein family have distinct amino acid sequences, they share structural similarities. Of particularly interest is the not yet fully understood self‐assembly ability of SCGBs, which arise from covalent disulfide dimerization and non‐covalent oligomerization. Recently, SCGB3A2 has attracted attention for its singular expression profile in airways. However, the knowledge on SCGB3A2 (patho)physiology derives exclusively from in vivo and complex ex vivo mixtures, which hampers characterization of the mechanisms driving SCGB3A2 structural behavior. Herein, we document the chemical synthesis of SCGB3A2 in multi‐milligram quantities. Key to access both monomeric and homodimeric SCGB3A2 analogues was the use of KAHA ligation and enabled masking of the cysteine residue. The synthetic proteins were used to investigate the SCGB3A2 self‐assembly profile, confirming their high propensity to dimerization even in the absence of the key Cys residue. [ABSTRACT FROM AUTHOR]

Hyperglycaemia is a life-threatening risk factor that occurs in both chronic and acute phases and has been linked to causing injury to many organs. Protein modification was triggered by hyperglycaemic stress, which resulted in pathogenic alterations such as impaired cellular function and tissue damage. Dysregulation in cellular function increases the condition associated with metabolic disorders, including cardiovascular diseases, nephropathy, retinopathy, and neuropathy. Hyperglycaemic stress also increases the proliferation of cancer cells. The major areas of experimental biomedical research have focused on the underlying mechanisms involved in the cellular signalling systems involved in diabetes-associated chronic hyperglycaemia. Reactive oxygen species and oxidative stress generated by hyperglycaemia modify many intracellular signalling pathways that result in insulin resistance and β-cell function degradation. The dysregulation of post translational modification in β cells is clinically associated with the development of diabetes mellitus and its associated diseases. This review will discuss the effect of hyperglycaemic stress on protein modification and the cellular signalling involved in it. The focus will be on the significant molecular changes associated with severe metabolic disorders. [ABSTRACT FROM AUTHOR]

This study aimed to modify plant protein mixture to improve their functionality and digestibility by limited hydrolysis. Soy protein isolate and corn zein were mixed at the ratio of 5:1 (w/w), followed by limited hydrolysis using papain from 15 to 30 min. The structural characteristics, in vitro digestibility, and functional properties were evaluated. Also, DPPH radical scavenging activity was determined. The results indicated that the molecular weight of different modified samples was largely reduced by limited hydrolysis, and the proportion of random coil was significantly increased. Furthermore, the solubility, foaming, emulsifying and water-holding capacity of hydrolyzed protein mixture were significantly improved, which were close to those of whey protein isolate. In vitro digestibility after 30-min limited hydrolysis was remarkably elevated. In addition, the hydrolyzed protein mixture exhibited a higher antioxidant activity than those of untreated proteins. Overall, limited hydrolysis of protein mixture led to improved digestibility, functionality and antioxidant activity.

Oxidative stress is widely recognized as a key mechanism in the development of hypertension. Under pathological conditions, such as in hypertension, oxidative stress leads to irreversible posttranslational modifications of proteins, which result in loss of protein function and cellular damage. We have previously documented physiological and morphological changes across various blood and bone marrow cell lineages, all of which exhibit elevated oxidative stress. While cytoskeletal changes in erythrocytes have been well characterized in hereditary diseases, this is the first study, to our knowledge, to investigate cytoskeletal reorganization in erythrocytes from hypertensive patients. To this end, we compared the expression patterns and subcellular distribution of key cytoskeletal proteins in erythrocytes from hypertensive individuals with those from normotensive subjects using Western blot, flow cytometry, and confocal microscopy. Our results revealed the presence of three erythrocyte subpopulations with differential expression of glycophorin A. The persistent oxidative environment in hypertensive patients causes dysregulation in the expression of glycophorin A, Band 3 protein, protein 4.1, and ankyrin, as well as the reorganization of spectrin. These alterations in protein expression and distribution suggest that oxidative stress in hypertensive individuals may induce structural modifications, ultimately impairing erythrocyte membrane elasticity and function.

Covalent modification of proteins at specific, predetermined sites is essential for advancing biological and biopharmaceutical applications. Site-selective labeling techniques for protein modification allow us to effectively track biological function, intracellular dynamics, and localization. Despite numerous reports on modifying target proteins with functional chemical probes, unique organic reactions that achieve site-selective integration without compromising native functional properties remain a significant challenge. In this review, we delve into site-selective protein modification using synthetic probes, highlighting both chemical and computational methodologies for chemo- and regioselective modifications of naturally occurring amino acids, as well as proximity-driven protein-selective chemical modifications. We also underline recent traceless affinity labeling strategies that involve exchange/cleavage reactions and catalyst tethering modifications. The rapid development of computational infrastructure and methods has made the bioconjugation of proteins more accessible, enabling precise predictions of structural changes due to protein modifications. Hence, we discuss bioconjugational computational approaches, including molecular dynamics and artificial intelligence, underscoring their potential applications in enhancing our understanding of cellular biology and addressing current challenges in the field.

Auxenochlorella pyrenoidosa is a microalga that stands out due to its high protein content. The objective of this work was to study the effect of high-pressure homogenization (HPH) on the recovery of proteins from A. pyrenoidosa and their application as functional emulsifiers. Untreated and HPH-treated (400–800 bar, 1 and 4 passes) aqueous cellular suspensions were incubated at 40 °C for 6 h. The aqueous extracts were collected, the proteins were precipitated at pH 3, and the Auxenochlorella pyrenoidosa protein concentrates (APPC) were lyophilized. Increasing HPH pressure and number of passes (400–800 bar, 1 and 4 passes) improved protein recovery yield up to 57%. Higher HPH pressures also reduced α-helix and β-sheet structures, exposing the hydrophobic protein core. This protein modification led to APPCs with increased oil-holding capacity (2.83 g oil/g APPC). The surface tension of APPC solutions reached a minimum value of 28.6 mN/m at an APPC concentration of 2% w/w. The APPCs from untreated and HPH-treated biomass were used to stabilize nanoemulsions (2–6% sunflower oil), comparing one-step homogenization (high-speed homogenization) with the two-step homogenization method (combining high-speed and high-pressure homogenization). The two-step method led to significantly smaller oil droplets with narrow size distribution, leading to stable nanoemulsions with improved resistance to centrifugation and heating–cooling cycles. Due to APPC’s great emulsifying properties, A. pyrenoidosa proteins have a promising potential for various applications such as delivery systems stabilization. Additionally, the low energy requirements, continuous processing capability, and scalability of HPH make it a suitable process for industrial applications.

Inspired by the enzyme lysyl oxidase, which selectively converts the side chain of lysine into allysine, an aldehyde‐containing post‐translational modification, we report herein the first chemical method for the synthesis of allysine by selective oxidation of dimethyl lysine. This approach is highly chemoselective for dimethyl lysine on proteins. We highlight the utility of this biomimetic approach for generating aldehydes in a variety of pharmaceutically active linear and cyclic peptides at a late stage for their diversification with various affinity and fluorescent tags. Notably, we utilized this approach for generating small‐molecule aldehydes from the corresponding tertiary amines. We further demonstrated the potential of this approach in generating cellular models for studying allysine‐associated diseases. [ABSTRACT FROM AUTHOR]

With a shift toward plant-based foods, pulses and grains belonging to the Fabaceae family, specifically pea, faba, or soybean varieties, have received attention recently as alternative protein sources due to their high protein content, nutritional profile, and emulsifying properties. Maillard reaction, caramelization of carbohydrates, or oxidation of lipids lead to generation and retention of bitter or beany off-flavors in these pulses and grains. Based on protein type, concentration, and functional groups of off-flavor compounds and their positioning, reversible or irreversible interactions between proteins and off-flavors occur that influence the taste of food products. The generation, retention, or interaction of off-flavors with these plant-based proteins can be controlled through different approaches. Bitter taste can be reduced during enzymatic hydrolysis or germination/sprouting due to the release of amino acids or masked during fermentation as a result of the generation of fruity or floral flavors. The pH variation or salt addition during protein extraction can modulate flavor-protein binding. Moreover, modification of proteins alters their surface hydrophobicity and charge, contributing to the reduction of off-flavors. Heating, exposure to electromagnetic field, or addition of biotic elicitors can decrease lipid oxidation and content of off-flavors with beany or grassy notes. The selected storage conditions and packaging materials also impact off-flavor retention and generation during storage. In light of the foregoing, this review article discusses recently followed processing approaches for controlling off-flavors in pea, faba, and soybeans as protein sources while providing an overview of recent studies performed on improving the sensory properties of food products produced from these alternative proteins. [ABSTRACT FROM AUTHOR]