Detection of Multiple Enzymes in Fermentation Broth Using Single PAGE Analysis.

Activity staining or zymography is a technique to detect enzymes based on their function/activity toward a specific substrate. Multiple enzyme-producing microbes secrete enzymes along with other proteins at varying time points during fermentation. The technique of zymography can be used to detect functionality of enzymes in complex protein/other enzyme mixtures. The protein bands corresponding to specific enzyme among other enzymes/proteins can be located by polyacrylamide gel electrophoresis (PAGE) followed by zymogram analysis. This can be employed to locate the secretion pattern of protein/enzyme from intracellular region to extracellular medium. Here we describe simple method for detection and cellular localization of esterases and protease secreted by single microbial strain in one PAGE gel.

Quantitative study on the effect of calcium and magnesium palmitate on the formation of Pseudomonas aeruginosa biofilm.

Calcium palmitate and magnesium palmitate (which are major constituents of waste water) are insoluble precipitates that accumulate in bodies of water. This leads to the formation of biofilms because bacterial cells can use these fatty acid salts as a carbon source. It is important to study the formation of biofilms because they cause corrosion of pipelines and water contamination. In this study, the effect of calcium palmitate and magnesium palmitate on Pseudomonas aeruginosa biofilm formation has been evaluated. In the presence of calcium palmitate, the biofilm biomass, extracellular polysaccharide, and adhesion force were 3.45 ± 0.06 (A590), 1810 ± 47 µg, and 14.5 ± 0.9 nN, respectively. In the presence of magnesium palmitate, the biofilm biomass, extracellular polysaccharide, and adhesion force were 2.72 ± 0.03 (A590), 1370 ± 56 µg, and 8.0 ± 0.2 nN, respectively. The results suggest that biofilm biomass, extracellular polysaccharide, and adhesion force were higher in the presence of calcium palmitate.

A quantitative study on the formation of Pseudomonas aeruginosa biofilm.

Biofilms are bacterial cells in a matrix of extracellular polymeric substance. The formation of biofilm depends on the microenvironment. In this study, the effect of temperature on Pseudomonas aeruginosa biofilm formation was evaluated with respect to three parameters-the mass of biofilm formed, the production of extracellular polysaccharide and the adhesion force. The results indicate that biofilm biomass (2.8, A590), extracellular polysaccharide production (1240 ± 40 µg) and adhesion force (10.8 ± 0.2 nN) were highest at 37°C. The results also suggest that biofilms formed at 37°C would have a higher mechanical stability (than biofilms grown at 28, 33 and 42°C).

Nanoscale investigation on Pseudomonas aeruginosa biofilm formed on porous silicon using atomic force microscopy.

Colonization of surfaces by bacterial cells results in the formation of biofilms. There is a need to study the factors that are important for formation of biofilms since biofilms have been implicated in the failure of semiconductor devices and implants. In the present study, the adhesion force of biofilms (formed by Pseudomonas aeruginosa) on porous silicon substrates of varying surface roughness was quantified using atomic force microscopy (AFM). The experiments were carried out to quantify the effect of surface roughness on the adhesion force of biofilm. The results show that the adhesion force increased from 1.5 ± 0.5 to 13.2 ± 0.9 nN with increase in the surface roughness of silicon substrate. The results suggest that the adhesion force of biofilm is affected by surface roughness of substrate.

Resonance Energy Transfer between protein and rhamnolipid capped ZnS quantum dots: application in in-gel staining of proteins.

The interaction of proteins with quantum dots is an interesting field of research. These interactions occur at the nanoscale. We have probed the interaction of Bovine Serum Albumin (BSA) and Candida rugosa lipase (CRL) with rhamnolipid capped ZnS (RhlZnSQDs) using absorption and fluorescence spectroscopy. Optical studies on mixtures of RhlZnSQDs and proteins resulted in Förster's Resonance Energy Transfer (FRET) from proteins to QDs. This phenomenon has been exploited to detect proteins in agarose gel electrophoresis. The activity of the CRL was unaffected on the addition of QDs as revealed by zymography.