Water-soluble Proteins Research Articles

Exacerbated interfacial impacts of nanoplastics and 6:2 chlorinated polyfluorinated ether sulfonate by natural organic matter in adult zebrafish: Evidence through histopathology, gut microbiota, and transcriptomic analysis

Nanoplastics (NPs) interact with cooccurring chemicals and natural organic matter (NOM) in the environment, forming complexes that can change their bioavailability and interfacial toxicity in aquatic organisms. This study aims to elucidate the single and combined impacts of 21-day chronic exposure to low levels of polystyrene NPs (size 80 nm) at 1 mg/L and 6:2 chlorinated polyfluorinated ether sulfonate (Cl-PFAES or F53B) at 200 μg/L in the presence and absence of NOM (humic acid-HA and bovine serum albumin-BSA at 10 mg/L) in adult zebrafish (Danio rerio). Our findings through multiple bioassays, revealed that the mixture group (M), comprising of NPs, F53B, HA, and BSA, caused a higher level of toxicity compared to the single NPs (AN), single F53B (AF), and combined NPs+F53B (ANF) groups. The mixture exposure caused the highest level of vacuolization and nuclear condensation in hepatocytes, and most of the intestinal villi were fused and highly reduced in villi length and crypt depth. Further, the T-AOC levels were significantly lower (p < 0.05), while the MDA levels in the liver and intestine were significantly higher (p < 0.05) in the M group with downregulation of nfkbiaa, while upregulation of prkcda, csf1ra, and il1b apoptosis genes in the liver. Pairwise comparison of gut microbiota showed significantly higher (p < 0.05) abundances of various genera in the M group, including Gordonia, Methylobacterium, Tundrisphaera, GKS98, Pedomicrobium, Clostridium, Candidatus and Anaerobacillus, as well as higher abundance of genera including pathogenic strains, while control group showed higher abundance of probiotic genus ZOR0006 than exposed group (p < 0.01). The transcriptomic analysis revealed highest number of DEGs in the M group (2815), followed by the AN group (506) and ANF group (206) with the activation of relaxin signaling pathway-RSP (slc9a1, slc9a2) and AMP-activated protein kinase (AMPK) pathway (plin1), and suppression of the toll-like receptor (TLR) pathway (tlr4a, tlr2, tlr1), cytokine-cytokine receptor interaction (CCRI) pathway (tnfb, il21r1, il21, ifng1), and peroxisome proliferator-activated receptors (PPAR) pathway (pfkfb3). Overall, toxicity in the M group was higher, indicating that the HA and BSA elevated the interfacial impacts of NPs and F53B in adult zebrafish after chronic environmentally relevant exposure, implying the revisitation of the critical interaction of NOM with co-occurring chemicals and associated impacts

Physiological and biochemical studies of young sterlet sturgeon (Acipenser ruthenus L.) in the lower reaches of the Volga river

In conditions of a decrease in the number of rolled sturgeon larvae in the lower reaches of the Volga River, an urgent issue is the study of the physiological and biochemical parameters of the fish organism. The purpose of this work is to conduct physiological and biochemical studies of juvenile sterlet (Acipenser ruthenus L, 1758). The study of fish was carried out using the methods of classical histology and biochemistry. The degree of damage to the gills and liver was assessed on a scale from 1 to 5 points. The content of total lipids and water-soluble protein in the muscle tissue of sterlet fingerlings was studied as biochemical parameters. The morphological picture of the gills of 80% of the fish corresponded to a weakly pronounced character of changes (2 points), while in others it was characterized by the presence of a borderline state between weakly pronounced changes and moderate injuries (2–3 points). Six types of disorders were found in the gills: hyperplasia of the respiratory epithelium of lamellae, dilation of the blood vessel of the filament, goblet-shaped swellings of the apical part of the filament, invasions, expansion of the apical part of the lamellae, proliferation of connective tissue of lamellae. In the liver of 80% of fish, a borderline state was recorded between mild changes and moderate damage (2–3 points), in 13.3% of individuals the liver had mild damage (2 points), in 6.7% there were medium damage (3 points). Four types of disorders were found in the liver of juvenile sterlet: fullness of the liver capillaries, pronounced venous fullness, cavities, capillary dilation. In the muscle tissue of fingerlings, the concentrations of total lipids were in the range of 1.39–5.28%, and water-soluble protein — 45.59–57.97 mg/g. The study of the physiological and biochemical parameters of juvenile sturgeon in the lower reaches of the Volga River is very important for the biology of the species and monitoring the natural reproduction of these valuable fish species.

Dynamic Observation of the Membrane Interaction Processes of β-Lactoglobulin by Time-Resolved Vacuum-Ultraviolet Circular Dichroism.

The elucidation of protein-membrane interactions is pivotal for comprehending the mechanisms underlying diverse biological phenomena and membrane-related diseases. In this investigation, vacuum-ultraviolet circular dichroism (VUVCD) spectroscopy, utilizing synchrotron radiation (SR), was employed to dynamically observe membrane interaction processes involving water-soluble proteins at the secondary-structure level. The study utilized a time-resolved (TR) T-shaped microfluidic cell, facilitating the rapid and efficient mixing of protein and membrane solutions. This system was instrumental in acquiring measurements of the time-resolved circular dichroism (TRCD) spectra of β-lactoglobulin (bLG) during its interaction with lysoDMPG micelles. The results indicate that bLG undergoes a β-α conformation change, leading to the formation of the membrane-interacting state (M-state), with structural alterations occurring in more than two steps. Global fitting analysis, employing biexponential functions with all of the TRCD spectral data sets, yielded two distinct rate constants (0.18 ± 0.01 and 0.06 ± 0.003/s) and revealed a unique spectrum corresponding to an intermediate state (I-state). Secondary-structure analysis of bLG in its native (N-, I-, and M-states) highlighted that structural changes from the N- to I-states predominantly occurred in the N- and C-terminal regions, which were prominently exposed to the membrane. Meanwhile, transitions from the I- to M-states extended into the inner barrel regions of bLG. Further examination of the physical properties of α-helical segments, such as effective charge and hydrophobicity, revealed that the N- to I- and I- to M-state transitions, which are ascribed to first- and second-rate constants, respectively, are primarily driven by electrostatic and hydrophobic interactions, respectively. These findings underscore the capability of the TR-VUVCD system as a robust tool for characterizing protein-membrane interactions at the molecular level.

Design of a water-soluble transmembrane receptor kinase with intact molecular function by QTY code

Membrane proteins are critical to biological processes and central to life sciences and modern medicine. However, membrane proteins are notoriously challenging to study, mainly owing to difficulties dictated by their highly hydrophobic nature. Previously, we reported QTY code, which is a simple method for designing water-soluble membrane proteins. Here, we apply QTY code to a transmembrane receptor, histidine kinase CpxA, to render it completely water-soluble. The designed CpxAQTY exhibits expected biophysical properties and highly preserved native molecular function, including the activities of (i) autokinase, (ii) phosphotransferase, (iii) phosphatase, and (iv) signaling receptor, involving a water-solubilized transmembrane domain. We probe the principles underlying the balance of structural stability and activity in the water-solubilized transmembrane domain. Computational approaches suggest that an extensive and dynamic hydrogen-bond network introduced by QTY code and its flexibility may play an important role. Our successful functional preservation further substantiates the robustness and comprehensiveness of QTY code.

A novel ACE inhibitory peptide from Pelodiscus sinensis Wiegmann meat water-soluble protein hydrolysate

Pelodiscus sinensis meat is a nutritional food and tonic with angiotensin-converting enzyme (ACE) inhibitory activities. To identify the bioactive substances responsible, several bioinformatics methods were integrated to enable a virtual screening for bioactive peptides in proteins identified within a water-soluble protein fraction of Pelodiscus sinensis meat by Shotgun proteomics. The peptides were generated from the identified proteins by in silico proteolysis using six proteases. A comparison of the numbers of proteins suitable for digestion with each enzyme and the iBAQ (intensity-based absolute quantification) values for these proteins revealed that bromelain and papain were the most suitable proteases for this sample. Next, the water solubility, toxicity, and ADMET (absorption/distribution/metabolism/excretion/toxicity) properties of these peptides were evaluated in silico. Finally, a novel ACE inhibitory peptide IEWEF with an IC50 value of 41.33 µM was identified. The activity of the synthesized peptide was verified in vitro, and it was shown to be a non-competitive ACE inhibitor. Molecular docking revealed that IEWEF could tightly bind to C-ACE, and N-ACE with energies less than 0 kJ mol−1, and the peptide IEWEF can form hydrogen bonds with C-ACE and N-ACE respectively. These results provide evidence that bioactive peptides in the water-soluble protein fraction account for (at least) some of the ACE inhibitory activities observed in Pelodiscus sinensis meat. Furthermore, our research provides a workflow for the efficient identification of novel ACE inhibitory peptides from complex protein mixtures.

Comparison between high-pressure steam and vinegar stir-frying on grinding characteristics and nutrient components of turtle shells: Difference between carapace and plastron

The nutritional differences between the carapace and plastron of turtle shells, and their grinding challenges, are not well understood. This study aimed to evaluate the nutritional differences between a turtle shell’s carapace and plastron, and the effects of high-pressure steam (HP) and vinegar stir-frying (VS) pretreatments. The carapace showed higher crude protein and fat but lower ash than the plastron. Both pretreatments reduced grinding energy and increased ash content while decreasing protein and amino acids levels. HP resulted in more water-soluble extract (15.13 %), protein (17.26 %), and peptide (20.77 %), whereas total free amino acids (FAAs: 545.62 mg/100 g) were increased by VS. Nutritional value was comparable between carapace and plastron. Additionally, in contrast to the conventional VS pretreatment approach, the HP method not only enhances the grinding process of turtle shells but also promotes the release of their water-soluble constituents. This advancement sets a solid foundation for future comprehensive studies aimed at exploring the nutritional potential of turtle shells.

Anti-SARS-CoV-2 Viral Activity of Sweet Potato Trypsin Inhibitor via Downregulation of TMPRSS2 Activity and ACE2 Expression In Vitro and In Vivo.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic. Known as COVID-19, it has affected billions of people worldwide, claiming millions of lives and posing a continuing threat to humanity. This is considered one of the most extensive pandemics ever recorded in human history, causing significant losses to both life and economies globally. However, the available evidence is currently insufficient to establish the effectiveness and safety of antiviral drugs or vaccines. The entry of the virus into host cells involves binding to angiotensin-converting enzyme 2 (ACE2), a cell surface receptor, via its spike protein. Meanwhile, transmembrane protease serine 2 (TMPRSS2), a host surface protease, cleaves and activates the virus's S protein, thus promoting viral infection. Plant protease inhibitors play a crucial role in protecting plants against insects and/or microorganisms. The major storage proteins in sweet potato roots include sweet potato trypsin inhibitor (SWTI), which accounts for approximately 60% of the total water-soluble protein and has been found to possess a variety of health-promoting properties, including antioxidant, anti-inflammatory, ACE-inhibitory, and anticancer functions. Our study found that SWTI caused a significant reduction in the expression of the ACE2 and TMPRSS2 proteins, without any adverse effects on cells. Therefore, our findings suggest that the ACE2 and TMPRSS2 axis can be targeted via SWTI to potentially inhibit SARS-CoV-2 infection.

The combined effects of ultrasound and plasma-activated water on silkworm pupae:Physicochemical properties, microbiological diversity and ultrastructure

A novel technique was proposed for processing silkworm pupae by combining plasma- activated water (PAW) with ultrasound (US). The microbial diversity and quality characteristics of the silkworm pupae were also evaluated. The results of the microbial diversity analysis indicated that PAW combined with US treatment significantly reduced the relative abundance of Streptococcaceae, Leuconostocaceae, and Acetobacteraceae from 32%, 18% and 16% to 27%, 11% and 11%, respectively. Microstructural analysis demonstrated that the collapse of the internal structure of chitin in silkworm pupae facilitated the release of nutrients and flavour compounds including fatty acids, water-soluble proteins (WSP), amino acids, phenolics, and volatile compounds. Furthermore, the increase in antioxidant capacity and the decrease in catalase activity and malondialdehyde content confirmed the mechanism of quality change. These findings provide new insights into the possible mechanism of PAW combined with US to improve the quality of edible insects.

Stingless bee honey: Nutritional, physicochemical, phytochemical and antibacterial validation properties against wound bacterial isolates.

With the rise of AMR the management of wound infections are becoming a big challenge. This has been attributed to the fact that most wound bacterial isolates have been found to possess various virulence factors like enzymes, toxins & biofilms production. Therefore, need for discovery of new lead compounds is paramount as such factors make these microbes to be resistant to already existing arsenal of antibiotics or even the immune system. This study aimed at documenting the nutritional, physicochemical, phytochemical and antibacterial properties of stingless bee honey. Isolation and characterization of bacterial isolates from 34 samples obtained from wounds of outpatients and surgical wards of Nakuru County Referral Hospital, Kenya was done. Various bacterial isolates (43) were isolated Staphylococcus aureus (34.8%) being predominant, followed by Pseudomonas aeruginosa (27.9%), Klebsiella pneumoniae (23.3%) and Escherichia coli (14.0%). A total of 36 out of the total isolates were genotypically characterized using molecular techniques detecting the prevalence of the following virulence genes; 16 srRNA (756 bp), hla (229 bp), cnf1 (426 bp), cnf2 (543 bp), hlyA (1011 bp), rmpA (461 bp), lasL (600 bp), gyrB (411 bp), khe (77 bp) and magA (128 bp). An assessment of the in vitro antibacterial activity of 26 stingless bee honey samples collected from their cerumen egg-shaped pots in Marigat sub-County, Baringo County, Kenya was done. Antibacterial properties of the stingless bee honey was done with varying susceptibility patterns being observed at different concentrations of honey impregnated discs (10x104, 20x104, 50x104 and 75x104 ml μg/ ml) giving mean inhibition diameters of 18.23 ± 0.4 mm (Staphylococcus aureus), 17.49 ± 0.3 mm (Pseudomonas aeruginosa), 16.05 ± 0.6 mm (Klebsiella pneumoniae) and 10.19 ± 0.5 mm (Escherichia coli) with a mean range of 14.54 ± 2.0 mm to 17.58 ± 3 mm. Higher susceptibility to honey was recorded across all the bacterial isolates compared to conventional antibiotics while the mean MIC and MBC of the honey were recorded at 62.5 ml μg/ ml and 250 ml μg/ ml respectively. Control bacterial isolates Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, Klebsiella pneumoniae ATCC 27736 and Pseudomonas aeruginosa ATCC 27858 were used in the analysis. The stingless bee honey was found to be rich in various nutritive components like sugar (89.85 ± 5.07 g/100 g) and moisture (81.75 ± 10.35 mg/g) with a significant difference of P <0.05 as the main antibacterial components. Additionally, the stingless honey did possess water soluble vitamins, proteins and minerals of which potassium was the most dominant one. In regard to phytochemicals, on our preliminary analysis phenolic, flavonoid and carotenoid compounds were found to be present with phenolic compounds being the most dominant one. Stingless bee honey from Marigat, has antimicrobial properties which could be attributed to the rich phytochemicals it possesses and its physicochemical properties in addition to its high nutritive value.

Theory of 2D electronic spectroscopy of water soluble chlorophyll-binding protein (WSCP): Signatures of Chl b derivate.

We investigate how electronic excitations and subsequent dissipative dynamics in the water soluble chlorophyll-binding protein (WSCP) are connected to features in two-dimensional (2D) electronic spectra, thereby comparing results from our theoretical approach with experimental data from the literature. Our calculations rely on third-order response functions, which we derived from a second-order cumulant expansion of the dissipative dynamics involving the partial ordering prescription, assuming a fast vibrational relaxation in the potential energy surfaces of excitons. Depending on whether the WSCP complex containing a tetrameric arrangement of pigments composed of two dimers with weak excitonic coupling between them binds the chlorophyll variant Chl a or Chl b, the resulting linear absorption and circular dichroism spectra and particularly the 2D spectra exhibit substantial differences in line shapes. These differences between Chl a WSCP and Chl b WSCP cannot be explained by the slightly modified excitonic couplings within the two variants. In the case of Chl a WSCP, the assumption of equivalent dimer subunits facilitates a reproduction of substantial features from the experiment by the calculations. In contrast, for Chl b WSCP, we have to assume that the sample, in addition to Chl b dimers, contains a small but distinct fraction of chemically modified Chl b pigments. The existence of such Chl b derivates has been proposed by Pieper et al. [J. Phys. Chem. B 115, 4042 (2011)] based on low-temperature absorption and hole-burning spectroscopy. Here, we provide independent evidence.

Water-soluble compounds from Lignosus rhinocerus TM02® (xLr™) modulate ACE2 activity and inhibit its interaction with SARS-CoV-2 Spike-Protein

Water-soluble compounds from Lignosus rhinocerus TM02® (xLr™) modulate ACE2 activity and inhibit its interaction with SARS-CoV-2 Spike-Protein

Use of turbidimetry for determination of heat treatment intensity applied at pasteurization of milk

Express methods for estimating the intensity of heat treatment of milk are necessary in industry and in research work. For this reason, there are many ways to measure this parameter, which are based on different physical principles, including turbidimetric methods. The Harland &amp; Ashworth turbidimetric method has been developed for a long time, however, due to its high reliability and ease of implementation, it is still used in practice. However, this method has a drawback: it takes a long time to perform the analysis. In this regard, the aim of the work is to develop an express method for evaluating the thermal class of milk based on the principle of measuring the concentration of soluble whey proteins. The result is achieved through the use of a turbidimetric measurement method with optimized sample preparation parameters and parameters for measuring the optical density of a suspension of protein aggregates. The method is implemented as follows. The milk sample is mixed with 0.1 N acetate buffer (pH 4.6) in a ratio that allows to obtain a concentration of soluble milk whey proteins from 0.05% to 0.1%. Recommended dilution coefficients: 1:3 for samples of ultra-pasteurized milk and pasteurized milk with high heat treatment intensity; 1:7 for samples of pasteurized milk with low heat treatment intensity and 1:14 for raw milk samples. The solution is filtered on a membrane filter with a pore size of 0.45 microns. The resulting filtrate is mixed with 24% trichloroacetic acid (TCA) in a 1:1 ratio to coagulate soluble whey proteins and form protein aggregates. After holding for 5–10 minutes, the optical density of the suspension of protein aggregates is measured at a wavelength of 650 nm. The content of water-soluble whey proteins in the sample can be calculated according to the calibration curve. The developed method allows to obtain the measurement result in less time than the Harland &amp; Ashworth turbidimetric method.

Arbuscular mycorrhizal symbiosis modulates nitrogen uptake and assimilation to enhance drought tolerance of Populus cathayana

This study aims to investigate effects of arbuscular mycorrhizal fungi (AMF) inoculation on nitrogen (N) uptake and assimilation in Populus cathayana under drought stress (DS). Herein, we measured photosynthetic performance, antioxidant enzyme system, N level and N assimilation enzymes, proteins content and distribution, transcripts of genes associated with N uptake or transport in P. cathayana with AMF (AM) or without AMF (NM) under soil water limitation and adequate irrigation. Compared with NM-DS P. cathayana, the growth, gas exchange properties, antioxidant enzyme activities, total N content and the proportion of water-soluble and membrane-bound proteins in AM-DS P. cathayana were increased. Meanwhile, nitrate reductase (NR) activity, NO3− and NO2− concentrations in AM-DS P. cathayana were reduced, while NH4+ concentration, glutamine synthetase (GS) and glutamate synthetase (GOGAT) activities were elevated, indicating that AM symbiosis reduces NO3− assimilation while promoting NH4+ assimilation. Furthermore, the transcriptional levels of NH4+ transporter genes (PcAMT1-4 and PcAMT2-1) and NO3− transporter genes (PcNRT2-1 and PcNRT3-1) in AM-DS P. cathayana roots were significantly down-regulated, as well as NH4+ transporter genes (PcAMT1-6 and PcAMT4-3) in leaves. In AM P. cathayana roots, DS significantly up-regulated the transcriptional levels of RiCPSI and RiURE, the key N transport regulatory genes in AMF compared with adequate irrigation. These results indicated that AM N transport pathway play an essential role on N uptake and utilization in AM P. cathayana to cope with DS. Therefore, this research offers a novel perspective on how AM symbiosis enhances plant resilience to drought at aspect of N acquisition and assimilation.

The Effects of Artificial Diets on the Expression of Molecular Marker Genes Related to Honey Bee Health.

Honey bees are commonly used to study metabolic processes, yet the molecular mechanisms underlying nutrient transformation, particularly proteins and their effects on development, health, and diseases, still evoke varying opinions among researchers. To address this gap, we investigated the digestibility and transformation of water-soluble proteins from four artificial diets in long-lived honey bee populations (Apis mellifera ligustica), alongside their impact on metabolism and DWV relative expression ratio, using transcriptomic and protein quantification methods. Diet 2, characterized by its high protein content and digestibility, was selected for further analysis from the other studied diets. Subsequently, machine learning was employed to identify six diet-related molecular markers: SOD1, Trxr1, defensin2, JHAMT, TOR1, and vg. The expression levels of these markers were found to resemble those of honey bees who were fed with Diet 2 and bee bread, renowned as the best natural food. Notably, honey bees exhibiting chalkbrood symptoms (Control-N) responded differently to the diet, underscoring the unique nutritional effects on health-deficient bees. Additionally, we proposed a molecular model to elucidate the transition of long-lived honey bees from diapause to development, induced by nutrition. These findings carry implications for nutritional research and beekeeping, underscoring the vital role of honey bees in agriculture.

Structural bioinformatics studies of glutamate transporters and their AlphaFold2 predicted water-soluble QTY variants and uncovering the natural mutations of L->Q, I->T, F->Y and Q->L, T->I and Y->F.

Glutamate transporters play key roles in nervous physiology by modulating excitatory neurotransmitter levels, when malfunctioning, involving in a wide range of neurological and physiological disorders. However, integral transmembrane proteins including the glutamate transporters remain notoriously difficult to study, due to their localization within the cell membrane. Here we present the structural bioinformatics studies of glutamate transporters and their water-soluble variants generated through QTY-code, a protein design strategy based on systematic amino acid substitutions. These include 2 structures determined by X-ray crystallography, cryo-EM, and 6 predicted by AlphaFold2, and their predicted water-soluble QTY variants. In the native structures of glutamate transporters, transmembrane helices contain hydrophobic amino acids such as leucine (L), isoleucine (I), and phenylalanine (F). To design water-soluble variants, these hydrophobic amino acids are systematically replaced by hydrophilic amino acids, namely glutamine (Q), threonine (T) and tyrosine (Y). The QTY variants exhibited water-solubility, with four having identical isoelectric focusing points (pI) and the other four having very similar pI. We present the superposed structures of the native glutamate transporters and their water-soluble QTY variants. The superposed structures displayed remarkable similarity with RMSD 0.528Å-2.456Å, despite significant protein transmembrane sequence differences (41.1%->53.8%). Additionally, we examined the differences of hydrophobicity patches between the native glutamate transporters and their QTY variants. Upon closer inspection, we discovered multiple natural variations of L->Q, I->T, F->Y and Q->L, T->I, Y->F in these transporters. Some of these natural variations were benign and the remaining were reported in specific neurological disorders. We further investigated the characteristics of hydrophobic to hydrophilic substitutions in glutamate transporters, utilizing variant analysis and evolutionary profiling. Our structural bioinformatics studies not only provided insight into the differences between the hydrophobic helices and hydrophilic helices in the glutamate transporters, but they are also expected to stimulate further study of other water-soluble transmembrane proteins.

Comparison of Extraction Assays and Quantification of Protein from Ulva anandii (Cholorophycota)

Seaweeds contain many macronutrients including protein, therefore they can be utilized to fulfil the protein requirements of human beings. This research focused on extracting total protein in green seaweed Ulva anandii (Amjad et Shameel 1993), from the crude extracts, by using the trichloroacetic acid (TCA) and acetone precipitation methods, and the estimation of crude extract (water-soluble proteins), and those obtained from the two above-mentioned methods. The results indicate that the water-soluble proteins had the highest quantity (949.75µg/mL) followed by the TCA precipitation method (831µg/mL), while the acetone precipitation method had the least concentration of total protein (100 µg/mL). The study concludes that treatment with organic solvents lowers the quantity of protein extracted from U. anandii.

Membrane Interaction Characteristics of the RTX Toxins and the Cholesterol-Dependence of Their Cytolytic/Cytotoxic Activity.

RTX toxins are important virulence factors produced by a wide range of Gram-negative bacteria. They are secreted as water-soluble proteins that are able to bind to the host cell membrane and insert hydrophobic segments into the lipid bilayer that ultimately contribute to the formation of transmembrane pores. Ion diffusion through these pores leads then to cytotoxic and cytolytic effects on the hosts. Several reports have evidenced that the binding of several RTX toxins to the target cell membrane may take place through a high-affinity interaction with integrins of the β2 family that is highly expressed in immune cells of the myeloid lineage. However, at higher toxin doses, cytotoxicity by most RTX toxins has been observed also on β2-deficient cells in which toxin binding to the cell membrane has been proposed to occur through interaction with glycans of glycosylated lipids or proteins present in the membrane. More recently, cumulative pieces of evidence show that membrane cholesterol is essential for the mechanism of action of several RTX toxins. Here, we summarize the most important aspects of the RTX toxin interaction with the target cell membrane, including the cholesterol dependence, the recent identification in the sequences of several RTX toxins of linear motifs coined as the Cholesterol Recognition/interaction Amino acid Consensus (CRAC), and the reverse or mirror CARC motif, which is involved in the toxin-cholesterol interaction.

Effects of microgels fabricated by microfluidic on the stability, antioxidant, and immunoenhancing activities of aquatic protein

Aquatic products are considered a potential source of novel bioactive proteins, which are used as therapeutic drugs for the treatment of different diseases (such as oxidative stress, immunocompromised, and inflammation), as well as nutraceuticals and cosmetics. However, the physical and chemical properties of proteins are unstable, and they are easily denatured by the influence of external high temperature and polar pH during processing, resulting in the loss of their functional activity. Herein, Fenneropenaeus chinensis water-soluble protein (FCWP) and Lateolabrax japonicus water-soluble protein (LJWP) were encapsulated within spherical biopolymer microgels composed of pectin and chitosan produced by the microfluidic device. The encapsulated samples remained inside the microgels when they were exposed to upper gastrointestinal but were released when they were exposed to simulated colonic fluid due to the hydrolysis effect by enzymes secreted by the colonic microflora. The results showed that microgels improve the thermal stability of FCWP and LJWP due to the interaction between polysaccharides and proteins in the microgels. In addition, microgels encapsulation did not affect the antioxidant and immunoenhancing activities of FCWP and LJWP. In summary, these microgels are suitable for oral colon-specific delivery in functional foods and supplements.

In Situ Spatiotemporal SERS Profiling of Bacterial Quorum Sensing by Hierarchical Hydrophobic Plasmonic Arrays in Agar Medium.

A feedback inhibition effect of high autoinducer levels on metabolite secretion in Chromobacterium subtsugae (C. subtsugae) was evidenced by in situ spatiotemporal surface-enhanced Raman spectroscopy (SERS) profiling. The hierarchical hydrophobic plasmonic array in agar medium is structured by oil/water/oil (OL/W/OH) triphasic interfacial self-assembly. The hydrophobic layer acts as a "door curtain" to selectively permit adsorption of a quorum sensing (QS)-regulated fat-soluble metabolite, i.e., violacein (Vio), and significantly blocks nonspecific adsorption of water-soluble proteins, etc. The SERS profiling clearly evidences that the diffusion of N-hexanoyl-l-homoserine lactone (C6-HSL) in agar medium quickly triggers the initial synthesis of Vio in C. subtsugae CV026 but surprisingly inhibits the intrinsic synthesis of Vio in C. subtsugae ATCC31532. The latter negative response might be related to the VioS repressor of ATCC31532, which negatively controls violacein production without influencing the expression of the CviI/R QS system. Moreover, two sender-receiver systems are constructed by separately coculturing CV026 or ATCC31532 with Hafnia alvei H4 that secretes large amounts of C6-HSL. Expectedly, the cocultivation similarly triggers the initial synthesis of Vio in CV026 but seems to have a quite weak negative effect on the intrinsic synthesis in ATCC31532. In fact, the negative regulation in ATCC31532 might be affected by a diffusion-dependent concentration effect. The H4 growth and its secretion of C6-HSL are a slow and continuous process, thereby avoiding the gathering of local high concentrations. Overall, our study put forward an in situ SERS strategy as an alternative to traditional bioluminescent tools for highly sensitively analyzing the spatiotemporal communication and cooperation in live microbial colonies.

Identification of Residues Potentially Involved in Optical Shifts in the Water-Soluble Chlorophyll a-Binding Protein through Molecular Dynamics Simulations.

Reversible light and thermally induced spectral shifts are universally observed in a wide variety of pigment-protein complexes at temperatures ranging from cryogenic to ambient. In this paper, we employed large-scale molecular dynamics (MD) simulations of a prototypical pigment-protein complex to better understand these shifts at a molecular scale. Although multiple mechanisms have been proposed over the years, no verification of these proposals via MD simulations has thus far been performed; our work represents the first step in this direction. From simulations of the water-soluble chlorophyll-binding protein complex, we determined that rearrangements of long hydrogen bonds were unlikely to be the origin of the multiwell landscape features necessary to explain observed spectral shifts. We also assessed small motions of amino acid residues and identified side chain rotations of some of these residues as likely candidates for the origin of relevant multiwell landscape features. The protein free-energy landscapes associated with side chain rotations feature energy barriers of around 1100-1600 cm-1, in agreement with optical spectroscopy results, with the most promising residue type associated with experimental signatures being serine, which possesses a symmetric triple-well landscape and moment of inertia of a relevant magnitude.