Cell immobilization induces changes in the protein response of Escherichia coli K-12 to a cold shock

We have compared the protein expression of gel-entrapped Escherichia coli cells submitted to a cold shock at 4 degrees C with those of exponential- and stationary-phase free-floating counterparts. Autoradiograms of two-dimensional gel electrophoresis patterns of proteins radiolabeled with L-[35S]methionine were compared using computing scanning densitometry. The levels of 203 proteins synthesized during the temperature shift were significantly and reproducibly higher than those corresponding to synthesis at 37 degrees C. A principal component analysis (PCA) was performed on the synthesis levels of these 203 proteins in the different incubation conditions tested. This study showed that the protein response of immobilized cells after the cold shock was significantly different from those of exponential- and stationary-phase free-floating organisms. For instance, protein SSB was specifically overexpressed by shocked immobilized organisms. Such induction of specific molecular mechanisms in immobilized bacteria might explain the high resistance of sessile-like organisms to stresses.     

540—Evolution with time of the zero-current potential of a gold electrode in Escherichia coli cultures supplied with lipoic acid: Part I. Mathematical modeling and computer simulation

Zero-current potential measurements (gold electrode) are suitable for continuously following the oxidation—reduction reactions of exogenic lipoic acid during Escherichia coli bacterial growth. This paper relates to a mathematical modeling of the experimental time-courses of potential.First, an empirical mathematical relation was obtained in vitro (i.e. in a sterile culture broth) between the zero-current potential and the concentrations of electroactive species that coexist and prevail in vivo (i.e. during the cultures).Secondly, a system of simple kinetic equations was proposed to express the metabolic, physical or chemical processes responsible for the in vivo time evolutions of the concentrations of electroactive species from which the time evolution of the electrode potential during the cultures was obtained. Most of these equations have been standardized by direct measurements. Numerical values could be applied to the remaining parameters of the model by comparing the computer-simulated time-courses of potential with experimental potential—time signals.The model properly fitted the experimental reality. It substantiated a theoretical correlation between the time evolutions of potential and the reductive activity of cultures by means of growth parameters relative to the population of organisms and transport or consumption parameters relative to the bacterial cell.     

Adhesion of Yersinia ruckeri to fish farm materials: influence of cell and material surface properties

Two strains (an environmental strain and a reference one coming from a national culture collection) of Yersinia ruckeri, a fish pathogenic bacterium, are characterised according to the ability to adhere on wood, concrete, polyvinylchloride (PVC) and fibreglass, four materials commonly found in fish farms. The relationships between adherence, bacterial and support hydrophobicities and surface roughness are investigated. The results show that: (i) Y. ruckeri is strongly hydrophilic; (ii) the environmental strain exhibits a higher ability to adhere than the reference one; and (iii) for the two strains a strong correlation is observed between roughness amplitude (RA) of the support material and adhesion ability.     

Substituting Coomassie Brilliant Blue for bromophenol blue in two-dimensional electrophoresis buffers improves the resolution of focusing patterns

In a new area of postgenomics challenges, the optimization of protein identification has become a central goal in microbiochemistry. In this work, we demonstrate that the substitution of Coomassie Brilliant Blue for bromophenol blue in two-dimensional electrophoresis (2-DE) buffers improves the focusing of whole proteins from Pseudomonas aeruginosa. This improvement of focusing concerns more particularly basic proteins. This enhancement may be attributed to a better transfer from the first to the second dimension, which probably highlights an increase in the solubility of proteins in the IPG strips. Hence, the use of an efficient tracking dye in the 2-DE buffers may enlarge protein recovery on proteome maps.     

Potentiometric detection and study of bacterial activity

The potentiometric technique leads to practical and fundamental applications, for the moment limited to the field of public health. Further possible extensions, e.g. the control of microbial contaminations in liquid foods, would bring it closer to industrial preoccupations. As an analytical application of the technique to the biotechnological processes, the feasibility of bacterial sensors, based on the potentiometric detection of lipoic acid reduction, may be considered for monitoring the concentrations of various substrates of microbial metabolism.     

Protein patterns of gel-entrapped Escherichia coli cells differ from those of free-floating organisms

The two-dimensional electrophoretic patterns of cellular proteins from gel-entrapped Escherichia coli cells were compared to those of exponential- and stationary-phase free-floating organisms. The amounts of several proteins in immobilized cells were significantly different from those in free bacteria. Immobilized organisms rapidly produced a high level of dipeptide permease and a single-strand binding protein, and progressively accumulated an aldehyde dehydrogenase. Immobilization also induced a decrease in the levels of two proteins, i.e., the YFID protein and a DNA-binding, stationary-phase protein. The possible role of these proteins in the high resistance of immobilized bacteria to stresses is discussed.     

541—Evolution with time of the zero-current potential of a gold electrode in escherichia coli cultures supplied with lipoic acid: Part II. Properties of the model and their application to investigations on drug effects on bacterial activity

In agreement with the in vivo time-courses of the gold electrode zero-current potential, the computer-simulated potential-time curves could be conveniently described by simple geometric magnitudes. We have investigated the influence exerted by the theoretical parameters of the model on these magnitudes.The lag time necessary for the sudden shift of potential to appear at low (10^?5M) initial concentration of exogenic LA was decreased by simulating an increase of the reductive activity of cultures. The slope of the potential-time curves at low and high (2 × 10^?4M) LA initial concentration changed in opposition with the lag time.These properties have suggested comparative in vivo potential-time measurements in the presence of drugs affecting bacterial activity. From the experimental modifications of the potential-time curves, the theoretical parameter(s) affected by the drug could be identified and discussed in relation to known results of molecular and cellular biology.Some illustrative examples of such an application have been developed. The effect of insulin on the potenlial-time curves has been attributed to a hormonal action at the bacterial cell membrane level. Marked differences in the effects of two different antibiotics (ampicillin and lividomycin) on the time evolutions of potential have also been analyzed and discussed within the frame of the theory.     

Protein synthesis in Escherichia coli at 4 degrees C

Two-dimensional electrophoresis technology was used to investigate protein synthesis by the mesophilic bacterium Escherichia coli at low temperature. It was confirmed that protein synthesis in E. coli decreased strongly after a temperature downshift from 37 to 4 degrees C. After incubation for 150 min at 4 degrees C, however, the number of synthesized proteins represented 60% of the overall polypeptide number observed at 37 degrees C. Furthermore, the analysis of autoradiograms revealed the overexpression of 69 proteins by shocked bacteria, showing that the number of cold-induced proteins has been significantly underestimated so far.