The role of bacterial cell wall hydrophobicity in adhesion

In this study, the adhesion of bacteria differing in surface hydrophobicity was investigated. Cell wall hydrophobicity was measured as the contact angle of water on a bacterial layer collected on a microfilter. The contact angles ranged from 15 to 70 degrees. This method was compared with procedures based upon adhesion to hexadecane and with the partition of cells in a polyethylene glycol-dextran two-phase system. The results obtained with these three methods agreed reasonably well. The adhesion of 16 bacterial strains was measured on sulfated polystyrene as the solid phase. These experiments showed that hydrophobic cells adhered to a greater extent than hydrophilic cells. The extent of adhesion correlated well with the measured contact angles (linear regression coefficient, 0.8).     

Interfacial thermodynamics of protein adsorption, ion co-adsorption and ion binding in solution. II. Model interpretation of ion exchange in lysozyme chromatography

In this paper we present a model for the ion exchange effects in protein adsorption. The model is applied to chromatography of lysozyme on strong cation exchanger ‘mono S’. The experimental and general thermodynamic aspects have been discussed in Part 1, the preceding paper. The main modelling assumptions are (i) the charge regulation is confined to the small layer of contact between adsorbed protein and exchanger surface, (ii) the contact layer as a whole is electroneutral and (iii) the number of protein acid/base groups and exchanger surface acid groups which participate in the ion exchange is proportional to the area of the contact layer. The model is fitted to the experimental data by adjustment of only two or three parameters. The experimental co-adsorption numbers are very well reproduced. A few conspicuous features emerge: (i) the number of protein acid/base groups and exchanger surface acid groups in the contact layer varies with the medium conditions, such that the number is higher when the interaction between protein and exchanger surface is stronger. (ii) There is indirect evidence for structural alterations in the upper layers of the exchanger surface: the adsorbed protein is probably partly ‘buried’ in the surface.     

The adsorption-induced secondary structure of beta-casein and of distinct parts of its sequence in relation to foam and emulsion properties

Changes in the secondary structure upon adsorption of beta-casein (betaCN) and of distinct parts of its sequence were investigated by far-ultraviolet circular dichroism in order to find suggested relationships with foam and emulsion-forming and -stabilising properties of the same protein/peptides. A teflon/water interface was used as a model system for foam and emulsion interfaces. The maximum surface loads of beta-casein and its derived peptides were investigated. The main secondary structure element of all samples in solution was the unordered random coil, but upon adsorption ordered structure, especially alpha-helix, was induced. At lower pH more ordered structure was induced, just as at lower ionic strength. Apparently, both hydrophobic and hydrophilic groups influence the change of secondary structure induced at a hydrophobic interface. The results suggest that the hydrophobic C-terminal half of betaCN accounted for the high maximum surface load on teflon, while the N-terminal half of betaCN seemed to be responsible for the secondary structure induction upon adsorption. A relation between the maximum surface load and the foam-stabilising properties was found, but an influence of the secondary structure properties on the foam and emulsion-forming and -stabilising properties was not observed.     

Overall charge and local charge density of pectin determines the enthalpic and entropic contributions to complexation with β-lactoglobulin

The complex formation between β-lactoglobulin and pectins of varying overall charge and local charge density were investigated. Isothermal titration calorimetry experiments were carried out to determine the enthalpic contribution to the complex formation at pH 4.25 and various ionic strengths. Complex formation was found to be an exothermic process for all conditions. Combination with previously published binding constants by Sperber et al. (Sperber, B. L. H. M.; Cohen Stuart, M. A.; Schols, H. A.; Voragen, A. G. J.; Norde, W. Biomacromolecules 2009, 10, 3246-3252) allows for the determination of the changes in the Gibbs energy and the change in entropy of the system upon complex formation between β-lactoglobulin and pectin. The local charge density of pectin is found to determine the balance between enthalpic and entropic contributions. For a high local charge density pectin, the main contribution to the Gibbs energy is of an enthalpic nature, supported by a favorable entropy effect due to the release of small counterions. A pectin with a low local charge density has a more even distribution of the enthalpic and entropic part to the change of the Gibbs energy. The enthalpic part is reduced due to the lower charge density, while the relative increase of the entropic contribution is thought to be caused by a change in the location of the binding place for pectin on the β-lactoglobulin molecule. The association of the hydrophobic methyl esters on pectin with an exposed hydrophobic region on β-lg results in the release of water molecules from the hydrophobic region and surrounding the methyl esters of the pectin molecule. An increase in the ionic strength decreases the enthalpic contribution due to the shielding of electrostatic attraction in favor of the entropic contribution, supporting the idea that the release of water molecules from hydrophobic areas plays a part in the complex formation.     

Structural features of a hyperthermostable endo-beta-1,3-glucanase in solution and adsorbed on "invisible" particles

Conformational characteristics and the adsorption behavior of endo-beta-1,3-glucanase from the hyperthermophilic microorganism Pyrococcus furiosus were studied by circular dichroism, steady-state and time-resolved fluorescence spectroscopy, and calorimetry in solution and in the adsorbed state. The adsorption isotherms were determined on two types of surfaces: hydrophobic Teflon and hydrophilic silica particles were specially designed so that they do not interact with light and therefore do not interfere with spectroscopic measurements. We present the most straightforward method to study structural features of adsorbed macromolecules in situ using common spectroscopic techniques. The enzyme was irreversibly adsorbed and immobilized in the adsorbed state even at high temperatures. Adsorption offered further stabilization to the heat-stable enzyme and in the case of adsorption on Teflon its denaturation temperature was measured at 133 degrees C, i.e., the highest experimentally determined for a protein. The maintenance of the active conformation and biological function particularly at high temperatures is important for applications in biocatalysis and biotechnology. With this study we also suggest that nature may employ adsorption as a complementary mode to maintain structural integrity of essential biomolecules at extreme conditions of temperature.     

Peptidylarginine deiminase 4 concentration,but not PADI4 polymorphisms,is associated with ICU mortality in septic shock patients

The objective of our study was to evaluate the association between peptidylarginine deiminase 4 (PAD4) concentration and its polymorphisms with mortality in patients with septic shock . We prospectively evaluated 175 patients aged over 18 years with septic shock upon intensive care unit (ICU) admission. However, 48 patients were excluded. Thus, 127 patients were enrolled in the study. At the time of the patients’ enrollment, demographic information was recorded. Blood samples were taken within the first 24 hours of the patient's admission to determine serum PAD4 concentrations and its polymorphism PADI4_89 [rs11203366], PADI4_94 [rs2240340] and PADI4_104 [rs1748033]. The mean age was 63.3 ± 15.2 years, 56.7% were male, PAD4 concentration was 4.62 (2.48‐6.20) ng/mL and the ICU mortality rate was 67.7%. The patients who died in the ICU had higher APACHE II and Sequential Organ Failure Assessment (SOFA) scores. In addition, PAD4 concentration was higher in patients who died during ICU stay. However, there were no differences regarding PADI4 polymorphisms and ICU mortality. In the logistic regression models, PAD4 concentrations were associated with ICU mortality when adjusted for APACHE II score and lactate (OR: 1.477; CI 95%: 1.186‐1.839; P < .001), and when adjusted for age, gender and APACHE II score (OR: 1.392; CI 95%: 1.145‐1.692; P < .001). In conclusion, PAD4 concentration, but not PADI4_89, PADI4_94 and PADI4_104 polymorphisms, is associated with ICU mortality in septic shock patients.     

X-ray photoelectron spectroscopy analysis of whole cells and isolated cell walls of gram-positive bacteria: comparison with biochemical analysis.

The surface chemical composition of whole cells and isolated cell walls of four coryneform bacteria and of a Bacillus brevis strain has been determined by X-ray photoelectron spectroscopy (XPS). The XPS data were converted into concentrations of model compounds: peptides, polysaccharides, and hydrocarbonlike compounds. The composition of the surface of B. brevis differed markedly from that of coryneforms: the peptide concentration was about twice higher in the former case, which is attributed to the presence of an S-layer at the cell surface; in contrast, the surface of coryneforms was rich in hydrocarbonlike compounds (about 40%), which was concomitant with a high water contact angle. The peptide surface concentration of the isolated cell walls of the five strains deduced from XPS data fitted well with the total peptide content determined by biochemical analysis, which supports the validity of XPS to determine the overall macromolecular composition of the bacterial cell surface. Compared to biochemical analysis of isolated cell walls, XPS analysis of whole cells provides information which concerns directly the cell surface (2- to 5-nm-thick layer) and is less subject to alteration via losses of cell wall constituents or contamination by intracellular compounds.     

The role of bacterial cell wall hydrophobicity in adhesion.

In this study, the adhesion of bacteria differing in surface hydrophobicity was investigated. Cell wall hydrophobicity was measured as the contact angle of water on a bacterial layer collected on a microfilter. The contact angles ranged from 15 to 70 degrees. This method was compared with procedures based upon adhesion to hexadecane and with the partition of cells in a polyethylene glycol-dextran two-phase system. The results obtained with these three methods agreed reasonably well. The adhesion of 16 bacterial strains was measured on sulfated polystyrene as the solid phase. These experiments showed that hydrophobic cells adhered to a greater extent than hydrophilic cells. The extent of adhesion correlated well with the measured contact angles (linear regression coefficient, 0.8).     

Electrophoretic mobility and hydrophobicity as a measured to predict the initial steps of bacterial adhesion.

The relationship between physiochemical surface parameters and adhesion of bacterial cells to negatively charged polystyrene was studied. Cell surface hydrophobicity and electrokinetic potential were determined by contact angle measurement and electrophoresis, respectively. Both parameters influence cell adhesion. The effect of the electrokinetic potential increases with decreasing hydrophobicity. Cell surface characteristics determining adhesion are influenced by growth conditions. At high growth rates, bacterial cells tend to become more hydrophobic. This fact can be of ecological significance for controlling the spread of bacteria throughout the environment.