Hydrophobic Protein Research Articles

Effect of glycosylated potato protein on the characteristics of gluten‐free dough and steamed bread

SummaryThe texture and sensory characteristics of gluten‐free steamed bread are highly dependent on the functional properties of protein, which can be improved by inducing the glycosylation reaction in protein‐polysaccharide mixtures. This study aimed to assess the impact of potato protein‐xanthan mixture (PX) and its glycosylated product (PXM) on the water state, rheological, specific volume and texture qualities of gluten‐free dough and steamed bread. The two mixtures were used to substitute for 0.5–2% of the flour in a trial gluten‐free steamed bread formulation. The specific volume (1.76 mL g−1) and hardness (13.40 N) of steamed bread with 2% PXM were significantly improved compared to those of the control (0.86 mL g−1, 19.17 N). Moreover, the addition of PX and PXM improved the water state, viscoelasticity and development height (Hm) of gluten‐free dough, especially PXM. The reason appeared to be related to that protein glycosylation modified the solubility, ζ‐potential, hydrophobicity and emulsion activity of potato protein. In conclusion, application of the glycosylated protein provides a potentially feasible approach to improve the quality of gluten‐free steamed bread.

Nitrogen regulates the synthesis of hydrophobic amino acids to improve protein structural and gel properties in common buckwheat

Nitrogen (N) fertilizer impacts the grain quality of common buckwheat, but the effects and regulatory mechanisms of N on various protein parameters of buckwheat are not fully understood. The purpose of this study was to investigate the particle morphology, structural and gel properties, and regulation mechanism of buckwheat protein under four N levels. The bulk density, surface hydrophobicity, particle size, and thermal properties of the buckwheat protein were maximized through the optimal N application (180 kg N/ha), further enhancing the thermal stability of the protein. N application increased the β-sheet content and reduced the random coil content. Appropriate N fertilizer input enhanced the tertiary structure stability and gel elasticity of buckwheat protein by promoting hydrophobic interactions, disulfide bonds, ionic bonds, storage modulus and loss modulus. The differentially expressed proteins induced by N are primarily enriched in small ribosomal subunit and ribosome, improving protein quality mainly by promoting the synthesis of hydrophobic amino acids. Future agriculture should pay attention to the hydrophobic amino acid content of buckwheat to effectively improve protein quality. This study further advances the application of buckwheat protein in the field of food processing and provides a theoretical basis for the extensive development and utilization of buckwheat protein.

Quinoa protein/polysaccharide electrostatic complex stabilized vegan high internal phase emulsions for 3D printing: Role of complex state and gelling-type polysaccharides

Rational selection of the complex state and polysaccharide type may enhance the performance of electrostatic complex stabilized high internal phase emulsions (HIPEs). Herein, quinoa proteins were extracted to form electrostatic complexes separately with three gelling-type polysaccharides to fabricate HIPEs. Results showed that the complexes in soluble state (pH 8.4–5.6) exhibited moderate size, high negative potential and enhanced protein hydrophobicity, and could achieve HIPEs with 84% oil phase upon acidification to pH 6 at low concentrations. Its excellent interfacial structure enhanced stability during heating, freeze-thawing and long-term storage, and exhibited promising 3D printing potential. Furthermore, the complexes formed by sulfated polysaccharide carrageenan had higher amphiphilicity than those formed by carboxylated polysaccharide pectin or sodium alginate, and their stabilized HIPE had preferable droplet size, stability and 3D printing resolution than its counterparts. This study may provide new insights into the performance enhancement of protein/polysaccharide electrostatic complex stabilized HIPEs.

Conformational flexibility and molecular weight influence the emulsification performance of pectin: A comparative study of sugar beet pectin and citrus pectin

Due to the complex composition and structure of plant polymers, a detailed investigation into the emulsification mechanism of pectin is still warranted. Herein, we aimed to clarify the relationship between the macromolecular structure (conformational flexibility and molecular weight (Mw) and the emulsification performance of pectin, which was obtained from two sources (sugar beet pectin (SBP) and citrus pectin (CP) and treated by hydrothermal depolymerization (dominated by acid hydrolysis), following which the overall flexibility was assessed using the persistence length (Lp)/Mw per unit contour length (ML) ratio. The results showed that depolymerization reduced the conformational flexibility of SBP (Lp/ML increased from 0.0169 to 0.0219 nm2 mol/g) and improved the flexibility of CP (Lp/ML decreased from 0.0551 to 0.0403 nm2 mol/g). Interfacial tension analysis revealed that the higher flexibility of pectin conferred superior surface activity, leading to decreased interfacial tension. Consequently, the fabricated emulsion was finer and more stable, with smaller changes in droplet size, as measured by a laser particle analyzer. The improved emulsification was attributed to the better exposure of hydrophobic proteins due to the conformational changes caused by the stretching and twisting of the more flexible polysaccharide chains in pectin, which facilitated higher interfacial adsorption of CP, increasing it from 43.4% to 61.8%. Moreover, the smaller interfacial concentration of the flexible pectins suggested that they had higher emulsification efficiency, in which the pectins might be densely packed at the interface to form a compact interfacial layer to stabilize the emulsion.

Diverse Roles of Protein Palmitoylation in Cancer Progression, Immunity, Stemness, and Beyond.

Protein S-palmitoylation, a type of post-translational modification, refers to the reversible process of attachment of a fatty acyl chain-a 16-carbon palmitate acid-to the specific cysteine residues on target proteins. By adding the lipid chain to proteins, it increases the hydrophobicity of proteins and modulates protein stability, interaction with effector proteins, subcellular localization, and membrane trafficking. Palmitoylation is catalyzed by a group of zinc finger DHHC-containing proteins (ZDHHCs), whereas depalmitoylation is catalyzed by a family of acyl-protein thioesterases. Increasing numbers of oncoproteins and tumor suppressors have been identified to be palmitoylated, and palmitoylation is essential for their functions. Understanding how palmitoylation influences the function of individual proteins, the physiological roles of palmitoylation, and how dysregulated palmitoylation leads to pathological consequences are important drivers of current research in this research field. Further, due to the critical roles in modifying functions of oncoproteins and tumor suppressors, targeting palmitoylation has been used as a candidate therapeutic strategy for cancer treatment. Here, based on recent literatures, we discuss the progress of investigating roles of palmitoylation in regulating cancer progression, immune responses against cancer, and cancer stem cell properties.

Stability of starch-proanthocyanidin complexes to in-vitro amylase digestion after hydrothermal processing.

Proanthocyanidins (PA) form poorly digestible complexes with starch. The study examined amylase degradation mechanism and hydrothermal stability of starch-PA complexes. Sorghum-derived PA was complexed with wheat starch, reconstituted into flour (10% gluten added) and processed into crackers and pancakes. In vitro digestion profile of the complexes and products were characterized. The starch-PA complexes retained more (34-84%) fragments with degree of polymerization (DP)>6,000 after 120min digestion than controls (0-21%). Debranching further revealed higher retention of DP 11 - 30 chains in the digested starch-PA complexes than controls, suggesting amylopectin complexation contributed to reduced starch digestion. Starch-PA complexes retained reduced digestibility (50-56% higher resistant starch vs controls) in the cracker, but not pancake model. However, removing gluten from the pancake formulation restored the reduced digestibility of the starch-PA complexes. The starch-PA complexes are stable to hydrothermal processing, but can be disrupted by hydrophobic gluten proteins under excess moisture conditions.

Improving the functionality of pea protein with laccase-catalyzed crosslinking mediated by chlorogenic acid

The influences of laccase-catalyzed crosslinking on the structural, emulsifying and gelling properties of pea protein with chlorogenic acid were intensively probed. Molecular weight analysis illustrated the formation of pea protein aggregates by laccase-induced polymerization in the presence of chlorogenic acid and the increase of incubation time facilitated the aggregation. Particle size of pea protein-laccase-chlorogenic acid progressively enhanced increasing incubation time. Laccase-induced polymerization possessed remarkable impacts on the secondary and tertiary structure of pea protein, confirmed by circular dichroism, and fluorescence spectroscopy. Surface hydrophobicity of pea protein appreciably enhanced with laccase-induced crosslinking due to the exposure of interior hydrophobic amino acid residues. Emulsifying activity, emulsifying capacity, gel strength, and water-holding capacity of pea protein can be considerably enhanced with laccase-catalyzed crosslinking with chlorogenic acid, suggesting excellent functionalities for pea protein were accomplished after being modified by laccase with chlorogenic acid. The obtained information will widen pea protein’s application in food systems.

A Research Journey: Over a Decade of Denaturing and Native-MS Analyses of Hydrophobic and Membrane Proteins in Amgen Therapeutic Discovery.

Membrane proteins and associated complexes currently comprise the majority of therapeutic targets and remain among the most challenging classes of proteins for analytical characterization. Through long-term strategic collaborations forged between industrial and academic research groups, there has been tremendous progress in advancing membrane protein mass spectrometry (MS) analytical methods and their concomitant application to Amgen therapeutic project progression. Herein, I will describe a detailed and personal account of how electrospray ionization (ESI) native mass spectrometry (nMS), ion mobility-MS (IM-MS), reversed phase liquid chromatographic mass spectrometry (RPLC-MS), high-throughput solid phase extraction mass spectrometry, and matrix-assisted laser desorption ionization mass spectrometry methods were developed, optimized, and validated within Amgen Research, and importantly, how these analytical methods were applied for membrane and hydrophobic protein analyses and ultimately therapeutic project support and progression. Additionally, I will discuss all the highly important and productive collaborative efforts, both internal Amgen and external academic, which were key in generating the samples, methods, and associated data described herein. I will also describe some early and previously unpublished nano-ESI (nESI) native-MS data from Amgen Research and the highly productive University of California Los Angeles (UCLA) collaboration. I will also present previously unpublished examples of real-life Amgen biotherapeutic membrane protein projects that were supported by all the MS (and IM) analytical techniques described herein. I will start by describing the initial nESI nMS experiments performed at Amgen in 2011 on empty nanodisc molecules, using a quadrupole time-of-flight MS, and how these experiments progressed on to the 15 Tesla Fourier transform ion cyclotron resonance MS at UCLA. Then described are monomeric and multimeric membrane protein data acquired in both nESI nMS and tandem-MS modes, using multiple methods of ion activation, resulting in dramatic spectral simplification. Also described is how we investigated the far less established and less published subject, that is denaturing RPLC-MS analysis of membrane proteins, and how we developed a highly robust and reproducible RPLC-MS method capable of effective separation of membrane proteins differing in only the presence or absence of an N-terminal post translational modification. Also described is the evolution of the aforementioned RPLC-MS method into a high-throughput solid phase extraction MS method. Finally, I will give my opinion on key developments and how the area of nMS of membrane proteins needs to evolve to a state where it can be applied within the biopharmaceutical research environment for routine therapeutic project support.

Delivery of Small-Molecule Drugs and Protein Drugs by Injectable Acid-Responsive Self-Assembled COF Hydrogels for Combinatorial Lung Cancer Treatment.

Covalent organic frameworks (COFs) have revealed enormous application prospects for cancer therapeutics recently, but their assembly systems face considerable challenges, such as the codelivery of hydrophobic and hydrophilic protein drugs with different physicochemical properties for in vivo delivery and release, as well as endosomal/lysosomal escape of protein drugs. To address these issues, we leveraged the high specific surface area, lipotropism, and structural tunability of boronate ester-linked COFs (COF-1) for the construction of advanced drug delivery systems. We first encapsulated the small-molecule drug doxorubicin (DOX) into a lipophilic COF (COF-1@DOX) and immobilized the functional protein drug ribonuclease A (RNase A) on the surface of the COF (RNase A-COF-1@DOX). We then created a novel composite delivery system (RNase A-COF-1@DOX gel) by cross-linking an albumin-oxygenated hydrogel (gel) network into the pores of COFs, allowing targeted codelivery of protein and small-molecule drugs in vivo. Using in-living body and multichannel fluorescence imaging, we analyzed the in vivo codelivery of protein and small-molecule drugs in a Lewis lung carcinoma (LLC) model. Finally, we applied the RNase A-COF-1@DOX gel to treat lung cancer in mice. This study paves an avenue for constructing COF-based drug delivery systems for lung cancer treatment and holds the potential to be extended to other types of cancer for more effective and targeted therapeutic treatments.

Glutaryl-CoA Dehydrogenase Misfolding in Glutaric Acidemia Type 1.

Glutaric acidemia type 1 (GA1) is a neurotoxic metabolic disorder due to glutaryl-CoA dehydrogenase (GCDH) deficiency. The high number of missense variants associated with the disease and their impact on GCDH activity suggest that disturbed protein conformation can affect the biochemical phenotype. We aimed to elucidate the molecular basis of protein loss of function in GA1 by performing a parallel analysis in a large panel of GCDH missense variants using different biochemical and biophysical methodologies. Thirteen GCDH variants were investigated in regard to protein stability, hydrophobicity, oligomerization, aggregation, and activity. An altered oligomerization, loss of protein stability and solubility, as well as an augmented susceptibility to aggregation were observed. GA1 variants led to a loss of enzymatic activity, particularly when present at the N-terminal domain. The reduced cellular activity was associated with loss of tetramerization. Our results also suggest a correlation between variant sequence location and cellular protein stability (p < 0.05), with a more pronounced loss of protein observed with variant proximity to the N-terminus. The broad panel of variant-mediated conformational changes of the GCDH protein supports the classification of GA1 as a protein-misfolding disorder. This work supports research toward new therapeutic strategies that target this molecular disease phenotype.

Modification of whey protein isolate by ultrasound-assisted pH shift for complexation with carboxymethylcellulose: Structure and interfacial properties

The application of whey protein isolate (WPI) is limited because of its compact spherical structure. In this study, ultrasound-assisted pH shift was employed to modify WPI for complexation with carboxymethylcellulose (CMC). The foaming and emulsifying properties of WPI/CMC complexes were investigated. The results demonstrate that the pretreatment of ultrasound-assisted pH 12 shift increased the content of free sulfhydryl groups from 16.5 μmol/g to 34.7 μmol/g and enhanced protein hydrophobicity from 311.4 to 370.6 (p < 0.05). Compared to the complexes formed by untreated WPI and CMC, the complexes pretreated with ultrasound-assisted pH 12 shift had a smaller size of 293.4 nm and a more uniform distribution. Furthermore, WPI/CMC complexes pretreated by ultrasound-assisted pH 12 shift exhibited higher emulsifying activity and emulsion stability index, which were increased by 8.9 % and 42.6 % respectively, in comparison with the control group (p < 0.05). A positive correlation was found between the surface hydrophobicity of WPI and emulsifying activity of WPI/CMC complexes. Ultrasound-assisted pH 2 shift improved the foaming capacity of complexes by 28.3 % over the control (p < 0.05). All the results indicate that the interfacial properties of WPI/CMC complexes can be improved significantly by the combination of pH shift and ultrasound.

Structural bioinformatics studies of bacterial outer membrane beta-barrel transporters and their AlphaFold2 predicted water-soluble QTY variants.

Beta-barrel outer membrane proteins (OMP) are integral components of Gram-negative bacteria, eukaryotic mitochondria, and chloroplasts. They play essential roles in various cellular processes including nutrient transport, membrane stability, host-pathogen interactions, antibiotic resistance and more. The advent of AlphaFold2 for accurate protein structure predictions transformed structural bioinformatic studies. We previously used a QTY code to convert hydrophobic alpha-helices to hydrophilic alpha-helices in over 50 membrane proteins with all alpha-helices. The QTY code systematically replaces hydrophobic leucine (L), isoleucine (I), valine (V), and phenylalanine (F) with hydrophilic glutamine (Q), threonine (T), and tyrosine (Y). We here present a structural bioinformatic analysis of five outer membrane beta-barrel proteins with known molecular structures, including a) BamA, b) Omp85 (also called Sam50), c) FecA, d) Tsx, and e) OmpC. We superposed the structures of five native beta-barrel outer membrane proteins and their AlphaFold2-predicted corresponding QTY variant structures. The superposed structures of OMPs and their QTY variants exhibit remarkable structural similarity, as evidenced by residue mean square distance (RMSD) values between 0.206Å to 0.414Å despite the replacement of at least 22% (Transmembrane variation) of the amino acids in the transmembrane regions. We also show that native outer membrane proteins and QTY variants have different hydrophobicity patches. Our study provides important insights into the differences between hydrophobic and hydrophilic beta-barrels and validates the QTY code for studying beta-barrel membrane proteins and perhaps other hydrophobic aggregated proteins. Our findings demonstrate that the QTY code can be used as a simple tool for designing hydrophobic proteins in various biological contexts.

Synergistic Antimicrobial Hybrid Bio-Surface Formed by Self-Assembled BSA Nanoarchitectures with Chitosan Oligosaccharide.

Innovation in green, convenient, and sustainable antimicrobial packaging materials for food is an inevitable trend to address global food waste challenges caused by microbial contamination. In this study, we developed a biogenic, hydrophobic, and antimicrobial protein network coating for food packaging. Experimental results show that disulfide bond breakage can induce the self-assembly of bovine albumin (BSA) into protein networks driven by hydrophobic interactions, and chitosan oligosaccharide (COS) with antimicrobial activity can be stably bound in this network by electrostatic interactions. The inherent antimicrobial activity of COS and the numerous hydrophobic regions on the surface of the BSA-network give the BSA@COS-network significant in vitro antimicrobial ability. More importantly, the BSA@COS-network coating can prolong the onset of spoilage of strawberries in various packaging materials by nearly 3-fold in storage. This study shows how surface functionalization via protein self-assembly is integrated with the biological functioning of natural antibacterial activity for advanced food packaging applications.

First identification of canine adenovirus 1 in mink and bioinformatics analysis of its 100 K protein.

Animal trade favors the spreading of emerging canine adenovirus 1 (CAdV-1) in mink. Because the 100K protein is not exposed to the viral surface at any stage, it can be used to differentiate the vaccine from wild virus infection. However, no related research has been conducted. This study aimed to find evidence of CAdV-1 in mink and predict the character of the 100K protein in the current circulating CAdV-1 strain of mink. In this experiment, the identification of CAdV-1, the phylogenetic tree, homology, and bioinformatics analysis of 100K were conducted. The results showed that the CAdV-1 was identified in the mink and that its Fiber was located in a separate branch. It was closely related to strains isolated from Norwegian Arctic fox and Red fox. 100K was located in a separate branch, which had the closest genetic relationship with skunks, porcupines, raccoons, and hedgehogs and a far genetic relationship with the strains in dogs. 100K protein is an unstable and hydrophobic protein. It had evidence of selective pressure and recombination, 1 glycosylation site, 48 phosphorylation sites, 60 dominant B cell epitopes, and 9 peptides of MHC-I and MHC-II. Its subcellular localization was mainly in the endoplasmic reticulum and mitochondria. The binding sites of 100K proteins were DBP proteins and 33K proteins. The stains in the mink were different from fox. The exploration of its genomic characteristics will provide us with a deeper understanding of the prevention of canine adenovirus.

Filamentous cyanobacteria and hydrophobic protein in extracellular polymeric substances facilitate algae—bacteria aggregation during partial nitrification

In algae—bacteria symbiotic wastewater treatment, the excellent settling performance of algae—bacteria aggregates is critical for biomass separation and recovery. Here, the composition of extracellular polymeric substances (EPS), microbial profiles, and functional genes of algae—bacteria aggregates were investigated at different solid retention times (SRTs) (10, 20, and 40 d) during partial nitrification in photo sequencing bioreactors (PSBRs). Results showed that SRTs greatly influenced the nitrogen transformation and the formation and morphological structure of algae—bacteria aggregates. The highest nitrite accumulation, the largest particle size (~1.54 mm) and the best settling performance were observed for the algae—bacteria aggregates in the PSBR with an SRT of 10 d, where the abundant occurrence of filamentous cyanobacteria with the highest ratio of chlorophyll a/b and the lowest EPS amount with the highest protein-to-polysaccharide ratio were observed. In particular, the EPS at 10 d of SRT contained a higher amount of protein-related hydrophobic groups and a lower ratio of α-helix/(β-sheet + random coil), indicating a looser protein structure, which might facilitate the formation and stabilization of algae—bacteria aggregates. Moreover, algal-bacterial aggregation greatly depended on the composition and evolution of filamentous cyanobacteria (unclassified _o__Oscillatoriales and Phormidium accounted for 56.29 % of the identified algae at SRT 10 d). The metagenomic analysis further revealed that functional genes related to amino acid metabolism (e.g., genes of phenylalanine, tyrosine, and tryptophan biosynthesis) were expressed at high levels within 10 d of SRT. Overall, this study demonstrates the influence of EPS structures and filamentous cyanobacteria on algae—bacteria aggregation and reveals the biological mechanisms driving photogranule structure and function.

Physiochemical characteristics, morphology, and lubricating properties of size-specific whey protein particles by acid or ion aggregation

To investigate the influence of particle characteristics on their lubricating capacity, microparticles of controlled size (~300, ~700, and ~1900 nm) were prepared from whey proteins using two different approaches: reducing the pH and increasing the calcium ion concentration. The physiochemical, morphological, and tribological properties of the two types of particles were determined. Both treatments pronouncedly decreased the absolute value of zeta-potential and surface hydrophobicity of whey proteins, with calcium ions showing a more severe effect on zeta-potential. The viscosity of the particle suspensions increased with particle size, and ion-induced samples showed higher viscosity than acid-induced ones. Morphology investigation revealed that particle aggregation and irregularity increased with particle size increase. Distinct lubricating behaviors were observed for the two particle types within different size ranges. Viscosity played a more important role in lubrication when the particle size was small, while particle characteristics became more dominant for large particles.

Effect of food-to-microorganisms ratio on aerobic granular sludge settleability: Microbial community, potential roles and sequential responses of extracellular proteins and polysaccharides

The food-to-microorganism ratio (F/M) is an important parameter in wastewater biotreatment that significantly affects the granulation and settleability of aerobic granular sludge (AGS). Hence, understanding the long-term effects and internal mechanisms of F/M on AGS settling performance is essential. This study investigated the relationship between F/M and the sludge volume index (SVI) within a range of 0.23–2.50 kgCOD/(kgMLVSS·d). Thiothrix and Candidatus_Competibacter were identified as two dominant bacterial genera influencing AGS settling performance. With F/M increased from 0.27 kgCOD/(kgMLVSS·d) to 1.53 kgCOD/(kgMLVSS·d), the abundance of Thiothrix significantly increased from 0.20% to 27.02%, and the hydrophobicity of extracellular proteins (PN) decreased, which collectively reduced AGS settling performance. However, under high-F/M conditions, the gel-like polysaccharides (PS) effectively retained the granular biomass by binding to the highly abundant Thiothrix (53.65%). The progressive increment in biomass led to a concomitant reduction in F/M, resulting in the recovery of AGS settleability. In addition, two-dimensional correlation infrared spectroscopy analysis revealed the preferential responses of PN and PS to the increase and decrease of F/M, and the content and characteristics of PN and PS played important roles in granular settling. The study provides insight into the microbial composition and the potential role of extracellular polymer substances in the AGS sedimentation behavior, offering valuable theoretical support for stable AGS operation.

Japanese encephalitis virus NS4B inhibits interferon beta production by targeting TLR3 and TRIF

Japanese encephalitis virus (JEV) is a flavivirus transmitted by mosquitoes, causing epidemics of encephalitis in humans and reproductive disorders in pigs. This virus is predominantly distributed in Asian countries and causes tens of thousands of infections in humans annually. Interferon (IFN) is an essential component of host defense against viral infection. Multiple studies have indicated that multifunctional nonstructural proteins of flaviviruses suppress the host IFN response via various strategies to facilitate viral replication. The flaviviruses encoded nonstructural protein 4B (NS4B) is a multifunctional hydrophobic nonstructural protein widely involved in viral replication, pathogenesis and host immune evasion. In this study, we demonstrated that NS4B of JEV suppressed the induction of IFN-β production, mainly through targeting the TLR3 and TRIF (a TIR domain-containing linker that induces IFN-β) proteins in the TLR3 pathway. In a dual-luciferase reporter assay, JEV NS4B significantly inhibited the activation of IFN-β promoter induced by TLR3 and simultaneously treated with poly (I:C). Moreover, NS4B also inhibited the activation of IFN-β promoter triggered by interferon regulatory factor 3 (IRF3)/5D or its upstream molecules in TLR3 signaling pathway. Furthermore, NS4B inhibited the phosphorylation of IRF3 under the stimulation of TLR3 and TRIF molecules. Mechanistically, JEV NS4B interacts with TLR3 and TRIF and confirmed by co-localization and co-immunoprecipitation assay, thereby inhibiting the activation of downstream sensors in the TLR3-mediated pathway. Overall, our results provide a novel mechanism by which JEV NS4B interferes with the host's antiviral response through targeting TLR3 receptor signaling pathway.

In vitro antibiofilm and bacteriostatic activity of diacerein against Enterococcus faecalis

Enterococcus faecalis is one of the main pathogens that causes hospital-acquired infections because it is intrinsically resistant to some antibiotics and often is capable of biofilm formation, which plays a critical role in resisting the external environment. Therefore, attacking biofilms is a potential therapeutic strategy for infections caused by E. faecalis. Current research indicates that diacerein used in the treatment of osteoarthritis showed antimicrobial activity on strains of gram-positive cocci in vitro. In this study, we tested the MICs of diacerein using the broth microdilution method, and successive susceptibility testing verified that E. faecalis is unlikely to develop resistance to diacerein. In addition, we obtained a strain of E. faecalis HE01 with strong biofilm-forming ability from an eye hospital environment and demonstrated that diacerein affected the biofilm development of HE01 in a dose-dependent manner. Then, we explored the mechanism by which diacerein inhibits biofilm formation through qRT-PCR, extracellular protein assays, hydrophobicity assays and transcriptomic analysis. The results showed that biofilm formation was inhibited at the initial adhesion stage by inhibition of the expression of the esp gene, synthesis of bacterial surface proteins and reduction in cell hydrophobicity. In addition, transcriptome analysis showed that diacerein not only inhibited bacterial growth by affecting the oxidative phosphorylation process and substance transport but also inhibited biofilm formation by affecting secondary metabolism, biosynthesis, the ribosome pathway and luxS expression. Thus, our findings provide compelling evidence for the substantial therapeutic potential of diacerein against E. faecalis biofilms.

Evaluation of ohmic heating modified soybean protein isolate structure and antioxidant film under different catechin concentrations

To produce edible films with excellent antioxidant properties, soy protein isolate (SPI) was pretreated using ohmic heating (OH) to create more catechin binding sites. Compared with untreated SPI, ohmic heating-pretreated SPI (OH-SPI) had a looser secondary structure, higher fluorescence intensity, more free sulfhydryl groups, higher hydrophobicity (H0), and lower protein particles, all of which facilitated covalent binding between SPI and catechin. The binding equivalent of OH-SPI and catechin increased from 128.90 to 237.43 nmol/mg protein. The above results proved that ohmic heating pretreatment benefited the covalent binding between SPI and catechin and the development of the antioxidant film. Scanning electron microscopy (SEM) showed that catechin-modified OH-SPI (OH-SPI-CC) films had a rough and aggregated structure. As the concentration of catechin increased, the mechanical properties of the film improved while the barrier properties slightly decreased. Fourier transform infrared (FTIR) and X-ray diffraction (XRD) confirmed that the cross-linking between catechin and OH-SPI was covalent. In application testing of OH-SPI-CC film, OH-SPI-CC film at a catechin concentration of 4.0 mg/mL (OH-SPI-CC-4) showed the best DPPH and ABTS free radical scavenging rate and the highest cumulative release in food simulators. Therefore, cross-linking catechin with OH-SPI was an effective method for preparing antioxidant films.