The synergistic activity of triclosan and ciprofloxacin on biofilms of Salmonella Typhimurium

Triclosan is a biocide whose wide use has raised a debate about the potential benefits vs. hazards of the incorporation of antimicrobials in consumer products. The purpose of the present study was to determine whether exposure of biofilms of Salmonella enterica serovar Typhimurium to triclosan influences the tolerance of the bacteria towards antibiotics such as ciprofloxacin and vice versa. A synergistic antibiofilm activity was observed when the biofilms were treated with triclosan before or together with ciprofloxacin, and an additive activity was observed with planktonic cells. For example 500 μg mL⁻¹ triclosan and 500 μg mL⁻¹ ciprofloxacin reduced the number of viable cells in the biofilm by 1.6 and 0.5 log, respectively. However, the sequential treatment of 500 μg mL⁻¹ triclosan followed by ciprofloxacin resulted in 4.8 log reduction. Combination indexes (CI) for biofilms treated with triclosan followed by ciprofloxacin were 0.7, 0.32 and 0.25 for reduction of 90%, 99% and 99.9%, respectively, indicating a synergism. For planktonic cells, CIs were 1±0.1, indicating an additive effect. Therefore, it was suggested that triclosan weakens the ability of biofilm-associated cells to survive exposure to ciprofloxacin in the biofilm, probably by improving the permeability or the activity of ciprofloxacin.     

Proinsulin C-peptide - A Consensus Statement

In recent years the physiological role of the proinsulin C-peptide has received increasing attention, focusing on the potential therapeutic value of C-peptide replacement in preventing and ameliorating type 1 diabetic complications. In order to consolidate these new data and to identify the immediate directions of C-peptide research and its clinical usefulness, an International Symposium was held in Detroit, Michigan, on October 20–21, 2000, under the auspices of the Wayne State University/Morris Hood Jr. Comprehensive Diabetes Center. In this communication, we review the cellular, physiological and clinical effects of C-peptide replacement in animal models and in patients with type 1 diabetes. Finally, recommendations are presented as to the most urgent studies that should be pursued to further establish the biological action of C-peptide and its therapeutic value.     

A Segmental Semi Markov Model for protein secondary structure prediction

Hidden Markov Models (HMMs) are practical tools which provide probabilistic base for protein secondary structure prediction. In these models, usually, only the information of the left hand side of an amino acid is considered. Accordingly, these models seem to be inefficient with respect to long range correlations. In this work we discuss a Segmental Semi Markov Model (SSMM) in which the information of both sides of amino acids are considered. It is assumed and seemed reasonable that the information on both sides of an amino acid can provide a suitable tool for measuring dependencies. We consider these dependencies by dividing them into shorter dependencies. Each of these dependency models can be applied for estimating the probability of segments in structural classes. Several conditional probabilities concerning dependency of an amino acid to the residues appeared on its both sides are considered. Based on these conditional probabilities a weighted model is obtained to calculate the probability of each segment in a structure. This results in 2.27% increase in prediction accuracy in comparison with the ordinary Segmental Semi Markov Models, SSMMs. We also compare the performance of our model with that of the Segmental Semi Markov Model introduced by Schmidler et al. [C.S. Schmidler, J.S. Liu, D.L. Brutlag, Bayesian segmentation of protein secondary structure, J. Comp. Biol. 7(1/2) (2000) 233-248]. The calculations show that the overall prediction accuracy of our model is higher than the SSMM introduced by Schmidler.     

Inactivation of glucose oxidase by diperoxovanadate-derived oxidants.

Inactivation of glucose oxidase occurred in the presence of bromide, vanadate, H(2)O(2), and phosphate (the bromide system), and this was prevented by NADH or phenol red, a bromine acceptor. Glucose oxidase present during the reaction between diperoxovanadate and a reduced form of vanadate, vanadyl (the vanadyl system), but not added after mixing the reactants, was inactivated, and this was accompanied by a loss of binding of the dye, Coomassie blue, to the protein. The transient intermediate of the type OVOOV(O(2)), known to form in these reactions and used in the oxidation of bromide ion and NADH, appears to be responsible for inactivating glucose oxidase. In both systems, the inactivation of the enzyme was prevented by histidine and DTT, known to quench singlet-oxygen. By direct measurement of 1270-nm emission of singlet-oxygen, its generation was demonstrated in the bromide system, and in the reaction of hypohalous acids with diperoxovanadate, but not in the vanadyl system. By themselves both hypohalous acids, HOCl and HOBr inactivated glucose oxidase, and their prior reaction with H(2)O(2) during which singlet-oxygen was released, protected the enzyme. The results provide support for possible oxidative inactivation of glucose oxidase by diperoxovanadate-derived oxidants.     

Site-directed mutagenesis of arginine-89 supports the role of its guanidino side-chain in substrate binding by Cephalosporium acremonium isopenicillin N synthase

Isopenicillin N synthase (IPNS) catalyses a key step in the penicillin and cephalosporin biosynthetic pathway which involves the oxidative cyclisation of the acyclic peptide delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV) to isopenicillin N. Based on crystallographic evidence from the Aspergillus nidulans IPNS crystal structure complexed with the substrate ACV (Roach et al. (1997) Nature 387, 827-830), we were able to provide mutational evidence for the critical involvement of the conserved R-X-S motif in ACV binding in IPNS. The crystal structure further implicated arginine-87 in the binding of the aminoadipyl portion of ACV. Thus, in this study, the site-directed mutagenesis of the corresponding arginine-89 in Cephalosporium acremonium IPNS (cIPNS) was performed to ascertain its role in cIPNS. Alteration of arginine-89 to five amino acids from different amino acid groups, namely lysine, serine, alanine, aspartate and leucine, was performed and no activity was detected in all the mutants obtained when enzyme bioassays were performed. Furthermore, the solubility of the mutants was considerably lower than the wild-type cIPNS after expression at 37 degrees C, but could be recovered when the expression temperature was lowered to 25 degrees C. This suggests that arginine-89 could be critical for the activity of cIPNS due to its involvement in ACV binding and the solubility of wild-type enzyme.