Molecular Docking of Potential Inhibitors of Broccoli Myrosinase.

Glucosinolates are secondary metabolites occurring in Brassicaceae plants whose hydrolysis may yield isothiocyanates, widely recognized as health-promoting compounds. Myrosinase catalyzes this conversion. The chemical mechanism involves an unstable intermediary (thiohydroxamate-O-sulfonate) that spontaneously decomposes into isothiocyanates or other non-bioactive compounds depending on pH and cofactors. At acidic pH, non-bioactive compounds such as nitriles and thiocyanates are formed, while at neutral pH isothiocyanates are obtained. Broccoli myrosinase has been poorly studied so far. Recently, its amino acidic sequence was elucidated, and a structural model was built. The aim of this work was to study the molecular interaction of broccoli myrosinase with different ligands at acidic pH to propose possible inhibitors that prevent formation of undesirable compounds at acidic pH, and that at neutral pH dissociate from the enzyme, allowing formation of isothiocyanates. The interaction between broccoli myrosinase and 40 ligands was studied by molecular docking simulations. Both the enzyme and each inhibitor were set at pH 3.0. Amygdaline and arbutin showed the highest affinity to broccoli myrosinase in this condition. The residues that stabilize the complexes agree with those that stabilize the substrate (Gln207, Glu429, Tyr352, and Ser433). Accordingly, amygdaline and arbutin would perform as competitive inhibitors of myrosinase at pH 3.0.

Development of a dynamic model for ventral hernia mesh repair.

INTRODUCTION: The adequate way of mesh fixation in laparoscopic ventral hernia repair is still subject to debate. So far, simulation has only been carried out in a static way, thereby omitting dynamic effects of coughing or vomiting. We developed a dynamic model of the anterior abdominal wall. MATERIALS AND METHODS: An aluminium cylinder was equipped with a pressure controlled, fluid-filled plastic bag, simulating the abdominal viscera. A computer-controlled system allowed the control of influx and efflux, thus creating pressure peaks of up to 200 mmHg to simulate coughing and 290 mmHg to simulate vomiting. We tested fixation with tacks (Absorbatack, Covidien Deutschland, Neustadt a. D., Germany). The model was controlled for the friction coefficient of the tissue against the mesh and the physiologic elasticity of the abdominal wall surrogate. RESULTS: The model was able to create pressure peaks equivalent to physiologic coughs or vomiting. Physiologic elasticity was thereby maintained. We could show that the friction coefficient is crucial to achieve a physiologic situation. The meshes showed a tendency to dislocate with an increasing number of coughs (Fig. 4). Nevertheless, when applied in a plain manner, the meshes withstood more cough cycles than when applied with a bulge as in laparoscopic surgery. CONCLUSIONS: The dynamic movement of the abdominal wall, the friction between tissue and mesh and the way of mesh application are crucial factors that have to be controlled for in simulation of ventral abdominal hernia closure. We could demonstrate that patient specific factors such as the frequency of coughing as well as the application technique influence the long term stability of the mesh.

Bridging with reduced overlap: fixation and peritoneal grip can prevent slippage of DIS class A meshes.

PURPOSE: Ventral hernia repair can be performed safely using meshes which are primarily stable upon dynamic intermittent straining (DIS) at recommended overlap. In specific clinical situations, e.g., at bony edges, bridging of the hernial orifice with reduced overlap might be necessary. To gain insight into the durability of various applications, two different meshes with the best tissue grip known so far were assessed. METHODS: The model uses dynamic intermittent strain and comprises the repetition of submaximal impacts delivered via a hydraulically driven plastic containment. Pig tissue simulates a ventral hernia with a standardized 5 cm defect. Commercially available meshes classified as primarily stable at recommended overlap were used to bridge this defect at recommended and reduced overlap. RESULTS: Using Parietex Progrip®, the peritoneum adds sufficient stability at least to a 2.5 cm overlap. Using Dynamesh Cicat®, four gluing spots with Glubran® are sufficient to stabilize a 3.75 cm overlap. A 2.5 cm overlap is stabilized with eight bonding spots Glubran® and 8 bonding spots combined with four sutures stabilize a 1.25 cm overlap. Here again, an intact peritoneum stabilizes the reconstruction significantly. CONCLUSIONS: Based on a pig tissue model, a total of 23 different conditions were tested. A DIS class A mesh can be easily stabilized bridging a 5 cm hernial orifice with reduced overlap. Caution must be exerted to extend these results to other DIS classes and larger hernial orifices. Further DIS investigations can improve the durability of hernia repair.

A theory of protein-resin interaction in hydrophobic interaction chromatography.

Docking simulations were performed in order to investigate surface area of interaction between several ribonucleases and a reduced model for the hydrophobic moiety used in Phenyl Sepharose using the program AutoDock 3.0. For each ribonucelase, 80 independent simulations with populations consisting of 100 random structures were performed and from these the most probable docked protein-ligand conformations were obtained. A new methodology was used to select the most probable conformations, based on qualitative and quantitative considerations. The interacting amino acids in each protein were identified. The average surface hydrophobicity of the interfacial zone (local hydrophobicity, LH) was determined. The LH showed a high correlation level (r2 = 0.99) with the "hydrophobic contact area" (HCA) experimentally determined for the different ribonucleases as well as with the dimensionless retention time (r2 = 0.90). This study allowed us to identify the zones on the protein surface most probably involved in protein retention in HIC, without tedious experimental work. Given the good correlation level obtained, this new methodology may constitute a novel approach that could be used to predict protein behavior in HIC.

The non-enzymatic microbicidal activity of lysozymes.

T4 lysozyme was thought to destroy bacteria by its muramidase activity. However, we demonstrate here that amphipathic helix stretches in the C-terminus of T4 lysozyme mediate its bactericidal and fungistatic activities. In heat-denatured T4 lysozyme, the enzymatic activity is completely abolished but unexpectedly, the antimicrobial functions remain preserved. Small synthetic peptides corresponding to amphipathic C-terminal domains of T4 lysozyme show a microbicidal activity. Its membrane disturbing activity was directly demonstrated for bacterial, fungal and plant cells but not in a hemolysis assay. Comparable results were obtained with hen egg white lysozyme. This opens up many new opportunities for optimization of lysozymes as antimicrobial agents in various applications by protein engineering.

Effect of surface hydrophobicity distribution on retention of ribonucleases in hydrophobic interaction chromatography.

The effect of surface hydrophobicity distribution of proteins on retention in hydrophobic interaction chromatography (HIC) was investigated. Average surface hydrophobicity as well as hydrophobic contact area between protein and matrix were estimated using a classical thermodynamic model. The applicability of the model to predict protein retention in HIC was investigated on ribonucleases with similar average surface hydrophobicity but different surface hydrophobicity distribution. It was shown experimentally that surface hydrophobicity distribution could have an important effect on protein retention in HIC. The parameter "hydrophobic contact area," which comes from the thermodynamic model, was able to represent well the protein retention in HIC with salt gradient elution. Location and size of the hydrophobic patches can therefore have an important effect on protein retention in HIC, and the hydrophobic contact area adequately describes this.

New approaches for predicting protein retention time in hydrophobic interaction chromatography.

Hydrophobic interaction chromatography (HIC) is an important technique for the purification of proteins. In this paper, we review three different approaches for predicting protein retention time in HIC, based either on a protein's structure or on its amino-acidic composition, and we have extended one of these approaches. The first approach correlates the protein retention time in HIC with the protein average surface hydrophobicity. This methodology is based on the protein three-dimensional structure data and considers the hydrophobic contribution of the exposed amino acid residues as a weighted average. The second approach, which we have extended, is based on the high correlation level between the average surface hydrophobicity of a protein's hydrophobic interacting zone and its retention time in HIC. Finally, a third approach carries out a prediction of the average surface hydrophobicity of a protein, using only its amino-acidic composition, without knowing its three-dimensional structure. These models would make it possible to test different operating conditions for the purification of a target protein by computer simulations, and thus make it easier to select the optimal conditions, contributing to the rational design and optimization of the process.

Effect of feeding strategy on Zymomonas mobilis CP4 fed-batch fermentations and mathematical modeling of the system.

In this work, the effect of the feeding strategy in Zymomonas mobilis CP4 fed-batch fermentations on the final biomass and ethanol concentrations was studied. Highest glucose yields to biomass (0.018 g/g) and to ethanol (0.188 g/g) were obtained in fed-batch fermentations carried out using different feeding rates with a glucose concentration in the feed equal to 100 g/l. Lower values (0.0102 g biomass/g glucose and 0.085 g ethanol/ g glucose) were obtained when glucose accumulated to levels higher than 60 g/l. On the other hand, the highest biomass (5 g/l) and ethanol (39 g/l) concentrations were obtained using a glucose concentration in the feed equal to 220 g/l and exponentially varied feeding rates. Experimental data were used to validate the mathematical model of the system. The prediction errors of the model are 0.39, 14.36 and 3.24 g/l for the biomass, glucose and ethanol concentrations, respectively. Due to the complex relationship for describing the specific growth rate, a fed-batch culture in which glucose concentration is constant would not optimize the process.