Bovine Submaxillary Mucin Research Articles

A thermodynamic theory of sorption in glassy polymers

Sorption of vapors by glassy polymers is known to have properties that are different from those typical for sorption by rubbery polymers or liquids. Despite an importance for many practical applications, this difference is still not fully understood. We developed a thermodynamic theory for description of penetrant activity in the glassy state. The presented approach considers the glass transition in terms of a second order phase transition and utilizes heat capacity changes for calculation of thermodynamic activity. The formalism developed here explains a deviation of penetrant activity in the glassy state from equilibrium values. Moreover, the sorption-desorption hysteresis can be explained in thermodynamic terms as well. We demonstrate that not only the heat capacity change of the polymer, but also the heat capacity change of the penetrant should be considered to understand the difference between three cases: sorption by equilibrium liquid, absorption by glass and desorption from glass.The theory presented here is tested on experimental data on water sorption by maltodextrin and bovine submaxillary gland mucin. A very good fitting of the experimental data for both sorption and desorption cases is demonstrated. The values of heat capacity increments of the components obtained by fitting the penetrant activity data can be used to get new insights into the non-equilibrium properties of glassy materials.

The ability of Neoparamoeba perurans to bind to and digest non-fish-derived mucin: Insights into the amoeba's mechanism of action to overcome gill mucus production.

Amoebic gill disease (AGD) is caused by the marine amoeba Neoparamoeba perurans, a facultative parasite. Despite the significant impact this disease has on production of Atlantic salmon worldwide, the mechanisms involved in host-parasite interaction remains unknown. Excessive gill mucus secretion is reported as a host defence mechanism to prevent microbial colonization in the gill epithelium. Despite this response, N. perurans still attaches and proliferates. The present study aimed to investigate the interaction between N. perurans and mucin, the most abundant component in mucus. An in vitro adhesion assay using bovine submaxillary mucin (BSM) demonstrated that amoeba binding to mucin-coated substrate was significantly higher than to the BSA control. This binding interaction is likely glycan-mediated as pre-incubation with galactose, galactosamine, N-acetylgalactosamine and fucose reduced mucin adhesion to control levels. The ability of N. perurans to secrete proteases that target mucin was also investigated. Protease activity was detected in the amoeba culture media in the presence of BSM, but not when protease inhibitor was added. Mucin degradation was visually assessed on protein gels. This study provides preliminary evidence that N. perurans has developed mechanisms to interact with and evade mucus by binding to mucin glycan receptors and secreting proteases with mucolytic activity.

Induction of ciliary orientation by matrix patterning and characterization of mucociliary transport

Impaired mucociliary clearance (MCC) is a key feature of many airway diseases, including asthma, bronchiectasis, chronic obstructive pulmonary disease, cystic fibrosis, and primary ciliary dyskinesia. To improve MCC and develop new treatments for these diseases requires a thorough understanding of how mucus concentration, mucus composition, and ciliary activity affect MCC, and how different therapeutics impact this process. Although differentiated cultures of human airway epithelial cells are useful for investigations of MCC, the extent of ciliary coordination in these cultures varies, and the mechanisms controlling ciliary orientation are not completely understood. By introducing a pattern of ridges and grooves into the underlying collagen substrate, we demonstrate for the first time, to our knowledge, that changes in the extracellular matrix can induce ciliary alignment. Remarkably, 90% of human airway epithelial cultures achieved continuous directional mucociliary transport (MCT) when grown on the patterned substrate. These cultures maintain transport for months, allowing carefully controlled investigations of MCC over a wide range of normal and pathological conditions. To characterize the system, we measured the transport of bovine submaxillary gland mucin (BSM) under several conditions. Transport of 5% BSM was significantly reduced compared with that of 2% BSM, and treatment of 5% BSM with the reducing agent tris(2-carboxyethyl)phosphine (TCEP) reduced viscosity and increased the rate of MCT by approximately twofold. Addition of a small amount of high-molecular-weight DNA increased mucus viscosity and reduced MCT by ∼75%, demonstrating that the composition of mucus, as well as the concentration, can have significant effects on MCT. Our results demonstrate that a simple patterning of the collagen substrate results in highly coordinated ciliated cultures that develop directional MCT, and can be used to investigate the mechanisms controlling the regulation of ciliary orientation. Furthermore, the results demonstrate that this method provides an improved system for studying the effects of mucus composition and therapeutic agents on MCC.

Development and optimisation of simulated salivary fluid for biorelevant oral cavity dissolution

Drug release within the oral cavity can be of paramount importance for formulations that are designed for specific purposes such as taste-masking, faster onset of therapeutic action, localization of treatment or avoidance of first-pass metabolism. Preclinical methods for assessment of dissolution in the oral cavity are necessary for design and development of these formulation but currently there is no consensus on what variables should be defined to achieve biorelevance in these tests. In this study, biorelevant simulated salivary fluids (SSFs) that can be uniformly applied for oral cavity dissolution testing were developed. Unstimulated saliva (US) SSF and stimulated saliva (SS) SSF were separately developed since the two states significantly differ. Physicochemical properties including pH, buffer capacity, surface tension and viscosity were assessed during development and optimised to mimic human saliva (HS). In order to account for the salivary proteins in HS, use of bovine submaxillary mucin (BSM) and porcine gastric mucin (PGM) in SSFs was evaluated. Following optimisation of the SSFs, biorelevance of the developed SSFs to HS was assessed by their comparative physicochemical properties as well as dissolution profiles of three diverse model compounds (sildenafil citrate, efavirenz, and caffeine) which showed comparable profiles between the SSFs and HS. This work addresses the lack of uniformed biorelevant dissolution media for oral cavity dissolution studies and provides a basis for standardised dissolution tests that provide consistency and harmonisation in future oral cavity dissolution studies. We envisage that this will have a positive impact on the development of new medicines that require functionality in the oral cavity.

Modulating the Bioactivity of Mucin Hydrogels with Crosslinking Architecture

AbstractHydrogels made of crosslinked macromolecules used in regenerative medicine technologies can be designed to affect the fate of surrounding cells and tissues in defined ways. Their function typically depends on the type and number of bioactive moieties such as receptor ligands present in the hydrogel. However, the detail in how such moieties are presented to cells can also be instrumental. In this work, how the crosslinking architecture of a hydrogel can affect its bioactivity is explored. It is shown that bovine submaxillary mucins, a highly glycosylated and immune‐modulating protein, exhibit strikingly different bioactivities whether they are crosslinked through their glycans or their protein domains. Both the susceptibility to enzymatic degradation and macrophage response are affected, while rheological properties and barrier to diffusion are mostly unaffected. The results suggest that crosslinking architecture affects the accessibility of the substrate to proteases and the pattern of sialic acid residues exposed to the macrophages. Thus, modulating the accessibility of binding sites through the choice of the crosslinking strategy appears as a useful parameter to tune the bioactivity of hydrogel‐based systems.

Viscoelastic particle focusing in human biofluids.

Saliva and blood plasma are non-Newtonian viscoelastic fluids that play essential roles in the transport of particulate matters (e.g., food and blood cells). However, whether the viscoelasticity of such biofluids alters the dynamics of suspended particles is still unknown. In this study, we report that under pressure-driven microflows of both human saliva and blood plasma, spherical particles laterally migrate and form a focused stream along the channel centerline by their viscoelastic properties. We observed that the particle focusing varied among samples on the basis of sampling times/donors, thereby demonstrating that the viscoelasticity of the human biofluids can be affected by their compositions. We showed that the particle focusing, observed in bovine submaxillary mucin solutions, intensified with the increase in mucin concentration. We expect that the findings from this study will contribute to the understanding of the physiological roles of viscoelasticity of human biofluids.

N-Glycan Modifications with Negative Charge in a Natural Polymer Mucin from Bovine Submaxillary Glands, and Their Structural Role.

Bovine submaxillary mucin (BSM) is a natural polymer used in biomaterial applications for its viscoelasticity, lubricity, biocompatibility, and biodegradability. N-glycans are important for mucin stability and function, but their structures have not been fully characterized, unlike that of O-glycans. In this study, BSM N-glycans were investigated using liquid chromatography-tandem mass spectrometry. The microheterogeneous structures of 32 N-glycans were identified, and the quantities (%) of each N-glycan relative to total N-glycans (100%) were obtained. The terminal N-acetylgalactosamines in 12 N-glycans (sum of relative quantities; 27.9%) were modified with mono- (10 glycans) and disulfations (2 glycans). Total concentration of all sulfated N-glycans was 6.1 pmol in BSM (20 µg), corresponding to 25.3% of all negatively charged glycans (sum of present N-glycans and reported O-glycans). No N-glycans with sialylated or phosphorylated forms were identified, and sulfate modification ions were the only negative charges in BSM N-glycans. Mucin structures, including sulfated N-glycans located in the hydrophobic terminal regions, were indicated. This is the first study to identify the structures and quantities of 12 sulfated N-glycans in natural mucins. These sulfations play important structural roles in hydration, viscoelasticity control, protection from bacterial sialidases, and polymer stabilization to support the functionality of BSM via electrostatic interactions.

Human neuraminidases have reduced activity towards modified sialic acids on glycoproteins

Multiple levels of diversity in sialic acid presentation can influence the substrate activity of sialosides for glycoside hydrolases. Few reports have investigated the specificity of human neuraminidase (hNEU) activity towards modified sialic acid residues that can occur on glycoproteins. Previously, we evaluated hNEU activity towards 9-O-acetylated sialic acid in glycolipid substrates and found that hNEU generally discriminated against 9-O-acetylated sialic acid over Neu5Ac. Here, we have investigated the substrate specificity of hNEU enzymes for a glycoprotein substrate (bovine submaxillary mucin) containing 9-O-acetylated and Neu5Gc residues. Using this model substrate, we observe a general trend for hNEU tolerance of Neu5Ac > Neu5Gc ≫ Neu5,9Ac2, consistent with our previous results with glycolipid substrates. These results expand our understanding of hNEU enzyme specificity and suggest that naturally occurring modifications of sialic acids can play a role in regulating hNEU activity.

Characterization of a novel O-acetyl sialic acid specific lectin from the hemolymph of the marine crab, Atergatis integerrimus (Lamarck, 1818)

An O-acetyl sialic acid specific lectin was purified from the hemolymph of the marine crab Atergatis integerrimus by affinity chromatography using BSM (Bovine Submaxillary Mucin) coupled to cyanogen bromide activated Sepharose 4B and biospecific adsorption using formalinized buffalo erythrocytes. The purified AiL (Atergatis integerrimus lectin) showed an 1218 fold increase in specific activity when compared to the crude hemolymph agglutinin. The lectin, on non - denaturing PAGE showed a single band of 216 kDa and when subjected to SDS - PAGE, the lectin resolved into three subunits of molecular weight 70, 72 and 74 kDa. Physico chemical characterization revealed the lectin as pH and temperature sensitive, calcium dependent and sensitive to calcium chelators. Based on the calcium dependency of the lectin, AiL could be classified as a C-type lectin. The purified lectin agglutinated buffalo erythrocytes with greater avidity and was inhibited by the glycoproteins BSM, thyroglobulin, fetuin, PSM, and sugars raffinose, trehalose, l - fucose, α - Lactose, melibiose and GluNAc suggesting the affinity of the lectin to sialic acid. Reduction in HA with asialo buffalo erythrocytes and HAI titer with desialylated BSM, confirms the sialic acid specificity of the lectin. The reduction in HAI following de - O - acetylation confirms the specificity of the lectin for O - acetyl sialic acid. FTIR analysis confirms the purified lectin as a glycoprotein with spectral bands corresponding to amide bands and saccharides. Thus this study paves way to assess the therapeutic application of this lectin that could be targeted to modified sialic acid moieties that are expressed on the malignant cells and pathogenic microbes and also deduce the crystal structure of the lectin.

Understanding the adsorption and potential tear film stability properties of recombinant human lubricin and bovine submaxillary mucins in an in vitro tear film model

The wetting and adsorption properties for two glycoproteins, recombinant human lubricin and bovine submaxillary mucins (BSM) were evaluated on hydrophilic and hydrophobic glass dome surfaces in a simplified in vitro tear film model. We show that both recombinant human lubricin (rh-lubricin) and BSM solutions render surfaces hydrophilic and when the fluid films reach 500 nm or less, the fluids resist evaporation-driven breakup through a volumetric flux across the surface, which we believe is due to evaporation-driven solutocapillary flows. rh-Lubricin was able to maintain a wet film without spontaneous breakup for longer periods of time than BSM at lower concentrations, which we attribute to differences in adsorption properties, measured by QCM-D, that result from surface charge and structural differences (confirmed by zeta potential, DLS, and SAXS measurements).

Structural and Quantitative Characterization of Mucin-Type O-Glycans and the Identification of O-Glycosylation Sites in Bovine Submaxillary Mucin.

Bovine submaxillary mucin (BSM) is a gel-forming glycoprotein polymer, and Ser/Thr-linked glycans (O-glycans) are important in regulating BSM’s viscoelasticity and polymerization. However, details of O-glycosylation have not been reported. This study investigates the structural and quantitative characteristics of O-glycans and identifies O-glycosylation sites in BSM using liquid chromatography–tandem mass spectrometry. The O-glycans (consisting of di- to octa-saccharides) and their quantities (%) relative to total O-glycans (100%; 1.1 pmol per 1 μg of BSM) were identified with 14 major (>1.0%), 12 minor (0.1%–1.0%), and eight trace (<0.1%) O-glycans, which were characterized based on their constituents (sialylation (14 O-glycans; 81.9%, sum of relative quantities of each glycan), non-sialylation (20; 18.1%), fucosylation (20; 17.5%), and terminal-galactosylation (6; 3.6%)) and six core structure types [Gal-GalNAc, Gal-(GlcNAc)GalNAc, GlcNAc-GalNAc, GlcNAc-(GlcNAc)GalNAc, and GalNAc-GalNAc]. O-glycosylation sites were identified using O-glycopeptides (bold underlined; 56SGETRTSVI, 259SHSSSGRSRTI, 272GSPSSVSSAEQI, 307RPSYGAL, 625QTLGPL, 728TMTTRTSVVV, and 1080RPEDNTAVA) obtained from proteolytic BSM; these sites are in the four domains of BSM. The gel-forming mucins share common domain structures and glycosylation patterns; these results could provide useful information on mucin-type O-glycans. This is the first study to characterize O-glycans and identify O-glycosylation sites in BSM.

Glyco-Modification of Mucin Hydrogels to Investigate Their Immune Activity.

Mucins are multifunctional glycosylated proteins that are increasingly investigated as building blocks of novel biomaterials. An attractive feature is their ability to modulate the immune response, in part by engaging with sialic acid binding receptors on immune cells. Once assembled into hydrogels, bovine submaxillary mucins (Muc gels) were shown to modulate the recruitment and activation of immune cells and avoid fibrous encapsulation in vivo. However, nothing is known about the early immune response to Muc gels. This study characterizes the response of macrophages, important orchestrators of the material-mediated immune response, over the first 7 days in contact with Muc gels. The role of mucin-bound sialic acid sugar residues was investigated by first enzymatically cleaving the sugar and then assembling the mucin variants into covalently cross-linked hydrogels with rheological and surface nanomechanical properties similar to nonmodified Muc gels. Results with THP-1 and human primary peripheral blood monocytes derived macrophages showed that Muc gels transiently activate the expression of both pro-inflammatory and anti-inflammatory cytokines and cell surface markers, for most makers with a maximum on the first day and loss of the effect after 7 days. The activation was sialic acid-dependent for a majority of the markers followed. The pattern of gene expression, protein expression, and functional measurements did not strictly correspond to M1 or M2 macrophage phenotypes. This study highlights the complex early events in macrophage activation in contact with mucin materials and the importance of sialic acid residues in such a response. The enzymatic glyco-modulation of Muc gels appears as a useful tool to help understand the biological functions of specific glycans on mucins which can further inform on their use in various biomedical applications.

Agglutination of plasma, hemocyanin, and separated hemocyanin from the hemolymph of the freshwater prawn Macrobrachium rosenbergii (De Man, 1879) (Decapoda: Caridea: Palaemonidae)

AbstractHemocyanin, the predominant protein in the hemolymph of crustaceans, was isolated from the plasma of the giant freshwater prawn Macrobrachium rosenbergii (De Man, 1879) and investigated, with its separated proteins MrHc1, 2, 3, and 4 (60, 114, 50, and 325 kDa, respectively) for hemagglutination (HA) activity. Hemocyanin against erythrocytes from human A, B, and O blood groups, mouse, rat sheep, water buffalo, and rabbit showed HA titer values higher than plasma. Of the four proteins isolated from hemocyanin, MrHc1 alone appeared to agglutinate the tested erythrocytes. Evaluated specific hemagglutination activity was higher for hemocyanin when compared to plasma. Sugar binding ability with N-acetylated sugars observed for plasma was restricted to N-acetyl neuraminic acid for hemocyanin and MrHc1. Inhibition of hemagglutination (HAI) by bovine submaxillary mucin (BSM) further defined the agglutination ability of hemocyanin and MrHc1. The hemocyanin protein MrHc1 on matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI–TOF/MS) and search in MASCOT database showed homology to hypothetical protein of Branchiostoma floridae Hubbs, 1922 (Cephalochordata). The results clearly indicated that the structure of the hemocyanin had evolved binding sites for sugar or sugar-linkages independently lacking homology with other hemocyanin or lectin-like proteins. The immunogenicity of this hemocyanin probably has biomedical applications as carrier adjuvants, which needs further investigation.

The antibiotic vancomycin induces complexation and aggregation of gastrointestinal and submaxillary mucins

Vancomycin, a branched tricyclic glycosylated peptide antibiotic, is a last-line defence against serious infections caused by staphylococci, enterococci and other Gram-positive bacteria. Orally-administered vancomycin is the drug of choice to treat pseudomembranous enterocolitis in the gastrointestinal tract. However, the risk of vancomycin-resistant enterococcal infection or colonization is significantly associated with oral vancomycin. Using the powerful matrix-free assay of co-sedimentation analytical ultracentrifugation, reinforced by dynamic light scattering and environmental scanning electron microscopy, and with porcine mucin as the model mucin system, this is the first study to demonstrate strong interactions between vancomycin and gastric and intestinal mucins, resulting in very large aggregates and depletion of macromolecular mucin and occurring at concentrations relevant to oral dosing. In the case of another mucin which has a much lower degree of glycosylation (~60%) – bovine submaxillary mucin - a weaker but still demonstrable interaction is observed. Our demonstration - for the first time - of complexation/depletion interactions for model mucin systems with vancomycin provides the basis for further study on the implications of complexation on glycopeptide transit in humans, antibiotic bioavailability for target inhibition, in situ generation of resistance and future development strategies for absorption of the antibiotic across the mucus barrier.

Identification, recombinant protein production, and functional analysis of a M60-like metallopeptidase, secreted by the liver fluke Opisthorchis viverrini

The carcinogenic liver fluke Opisthorchis viverrini (O. viverrini) is endemic in Thailand and neighboring countries including Laos PDR, Vietnam and Cambodia. Infections with O. viverrini lead to hepatobiliary abnormalities including bile duct cancer–cholangiocarcinoma (CCA). Despite decades of extensive studies, the underlying mechanisms of how this parasite survives in the bile duct and causes disease are still unclear. Therefore, this study aims to identify and characterize the most abundant protein secreted by the parasite. Proteomics and bioinformatics analysis revealed that the most abundant secretory protein is a metallopeptidase, named Ov-M60-like-1. This protein contains an N-terminal carbohydrate-binding domain and a C-terminal M60-like domain with a zinc metallopeptidase HEXXH motif. Further analysis by mass spectrometry revealed that Ov-M60-like-1 is N-glycosylated. Recombinant Ov-M60-like-1 (rOv-M60-like-1) expressed in Escherichia coli (E. coli) was able to digest bovine submaxillary mucin (BSM). The mucinase activity was inhibited by the ion chelating agent EDTA, confirming its metallopeptidase identity. The enzyme was active at temperatures ranging 25–37 °C in a broad pH range (pH 2–10). The identification of Ov-M60-like-1 mucinase as the major secretory protein of O. viverrini worms warrants further research into the role of this glycoprotein in the pathology induced by this carcinogenic worm.

Fabrication of eggshell membrane–based novel buccal mucosa–mimetic surface and mucoadhesion testing of chitosan oligosaccharide films

Abstract

N-glycans of bovine submaxillary mucin contain core-fucosylated and sulfated glycans but not sialylated glycans

Bovine submaxillary mucin (BSM) is a heavily-glycosylated macromolecular (approximately 4 MDa) protein and is used in various biomaterial applications in light of its high viscosity and biocompatibility, in addition to use as a biochemical substrate or inhibitor as a result of its abundant O-glycans. Although it has been reported that N-glycosylation provides stability of human mucins, most BSM research has been focused on its O-glycans, while N-glycans have not been reported to date. In this study, a common N-glycan core component was detected by monosaccharide analysis of BSM, and the structures of the N-glycans and their relative quantities were determined by liquid chromatography–tandem mass spectrometry. Seventeen N-glycans comprising ten complex-type [Fucose0~2Hexose3~4N-acetylhexosamine1~6Sulfate0~1; 61.1% (the sum of the relative quantities of each N-glycan out of the total N-glycans)], two high-mannose-type (Hexose5~6N-acetylhexosamine2; 12.0%), and five paucimannose type (Fucose0~1Hexose3~4N-acetylhexosamine2~3; 26.9%) were identified, but no hybrid-type or sialylated N-glycans were found. Additionally, these are less-branched structures compared to human mucins. Of these, ten glycans (77.2%), including two sulfated glycans (8.0%), were core fucosylated, which confer unique biological functions to glycoproteins. The N-glycosylation sites were identified from the analysis of glycopeptides from BSM. This study is the first confirmation of N-glycan attachment to BSM.

Submaxillary Mucin: its Effect on Aroma Release from Acidic Drinks and New Insight into the Effect of Aroma Compounds on its Macromolecular Integrity

Submaxillary mucin is a major component that defines the makeup and functionality of saliva. Understanding its structure and function during food intake is key to designing appropriate strategies for enhancing the delivery of flavour. In the present study, the hydrodynamic integrity of bovine submaxillary mucin was characterised under physiological and acidic conditions and it was shown to have a broad molecular weight distribution with species ranging from 100 kDa to over 2000 kDa, and a random coil type of conformation. A decrease in the pH of mucin appeared to result in aggregation and a broader molecular weight distribution, which was shown to correlate with a release of flavour compounds. Our study also provides indications that p-cresol may have an effect on the macromolecular integrity of mucin.

De-O-Acetylation of mucin-derived sialic acids by recombinant NanS-p esterases of Escherichia coli O157:H7 strain EDL933

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain EDL933 encodes the single chromosomal 9-O-acetylesterase NanS, and several copies of prophage-encoded 9-O-acetylesterases (NanS-p). These enzymes have recently been shown to cleave 5-N-acetyl-9-O-acetyl neuraminic acid (Neu5,9Ac2) to yield de-O-acetylated Neu5Ac, the latter of which may serve as a carbon and/or nitrogen source.In the current study, we investigated the NanS- and NanS-p-mediated digestion of synthetic O-acetylated neuraminic acids and bovine submaxillary glands mucin (BSM)-derived O-acetylneuraminic acids by high-performance thin-layer chromatography (HPTLC) and nano electrospray ionization mass spectrometry (nanoESI MS). Initial HPTLC analyses showed the expected activity of NanS and NanS-p variants for Neu5,9Ac2. However, all tested enzymes were unable to de-O-acetylate 5-N-acetyl-4-O-acetylneuraminic acid (Neu5,4 Ac2) in our test system. The nanoESI MS analysis of neuraminic acids after treatment of BSM with NanS-p gave evidence that NanS-p variants of EHEC O157:H7 strain EDL933 cleave off O-acetyl groups from mono-, di-, and tri-O-acetylated Neu5Ac and N-glycolylneuraminic acid (Neu5Gc), regardless of the carbon positions C7, C8 or C9 of the acetate esters. This enzyme activity leads to neuraminidase-accessible Neu5Ac and Neu5Gc on mucin glycans.Moreover, we could demonstrate by HPTLC analyses that recombinant Bacteroides thetaiotaomicron sialidase (BTSA-His) was able to cleave Neu5Ac and Neu5,9Ac2 from BSM and that the combination of BTSA-His with both NanS-His and NanS-p-His derivatives enhanced the release of de-O-acetylated core Neu5Ac and Neu5Gc from mammalian mucin O-glycans.Growth experiments with EHEC wildtype strain EDL933, its nanS and nanS/nanS-p1a-p7 mutant and exogenous BTSA-His in BSM demonstrated that the presence of BTSA-His enhanced growth of EDL933 and the nanS deletion mutant but not the nanS/nanS-p1a-p7 mutant.Thus, we hypothesize that the expression of sialic acid O-acetylesterases with a broad specificity could be an advantage in competition with the gut microbiota for nutrients and facilitate EHEC colonization in the human large intestine.

Interactions of Fly Ash Particles with Mucin and Serum Albumin.

Fly ash particles can contribute to haze and adverse health outcomes. In this study, two mucins, one from bovine submaxillary glands (bovine submaxillary mucin, BSM) and one from porcine stomach (porcine gastric mucin), as well as bovine serum albumin (BSA), which served as the physical barriers against foreign substances entering the tissues and the blood protein, respectively, were chosen as models for the investigations of the interactions between the proteins and the fly ash particles. Their adsorption behaviors were studied using spectroscopy and a quartz crystal microbalance with a dissipation monitor (QCM-D). The results indicated that the fly ash particles can induce the loosening of mucins and BSA, probably via the formation of complexes. Further, the secondary structure of proteins changed in the presence of fly ash particles. The α-helix content decreased with an increasing fly ash particle concentration. The addition of fly ash particles into protein solutions led to fluorescence quenching, which suggested that there were interactions between these particles and the mucins and BSA. The association constants ( Ka) for BSM and BSA were 5.35 and 4.18 L/g, respectively. Furthermore, the results of QCM-D analyses showed that the amount decreased on the mucin surface but increased slightly on the BSA surface, which indicated that the fly ash particles disrupted the mucin layer upon adsorption. These findings provide clear evidence of the interactions between the fly ash particles and the mucins and BSA, which can lead to structural changes. This study contributes to a better understanding of the interactions and adsorptions of atmospheric particulate pollutants with the proteins in the human body.