Bacterial cell extracts

Based on the previous publications, azo dye can be reduced by azoreductase-catalyzed reduction under anaerobic conditions. But still there is a speculation whether bacterial flavin reductases are responsible for the azo reductase activity observed with bacterial cell extracts. In a published report, it is reported that flavin reductases are indeed able to act as azo reductases [24]. Bacteria produce extracellular oxidative enzymes, which are relatively nonspecific enzymes catalyzing the oxidation of a variety of dyes. It was reported that so many diverse groups of bacteria play a role in decolorization. It has been also reported that mixed microbial community could reduce various azo dyes, and members of the y-proteabacteria and sulfate reducing bacteria (SRB) were found to be prominent members of mixed bacterial population by using molecular methods to determine the microbial population dynamics [1],... 

Fig. 6.4 Affinity electrophoresis of the recombinant phosphorylases from potato tuber.,3) A crude bacterial cell extract containing the type-L isozyme (lane 1), the chimeric enzyme (lane2), or the type-H isozyme (lane 3) was electrophoresed in 5% polyacrylamide gels supplemented with 0 (A), 50 (B), and 500 pg/ml (C) glycogen. After electrophoresis, the gels were stained for enzyme activity in KI-b solution. (Reproduced with permission from Goldsmith and Fletterick, Pure and Appl. Chern., 55(4), 583 (1983)).

Dissolution of preexisting collagen fibers in the immediate proximity of neoplastic cells is a well-recognized feature of active invasive growths. This has led many to postulate that tumor cells release collagenase, a specific proteolytic enzyme found in both mammalian and bacterial cells. Extracts of various animal and human tumors exhibit enhanced collagenase activity (104,131,270, 310, 319). 

We have found that it is possible to detect protein in bacterial cell extracts and nuclear extracts from mammalian cells. By using a specific antibody... 

Fig. 9 Normalised binding data obtained for purified human p52 DBD, bacterial cell extracts containing p52 DBD and mammalian cell extracts containing over-expressed full-length p52 on Codelink protein-DNA microarrays. The sequence GGGGTTCCCC (NF-kB14) was assigned a value of 1000 in each experiment. NF-kB18, NF-kB21 and NF-kB28 are classified as medium binders (Fig. 7) NF-kB30 is a low binder...

Fucumura, T. Hydrolysis of cyclic and liner oligomers of 6-aminocaproic acid by a bacterial cell extract. / of Biochemistry, 59 531-536 (1966). 

Lysozyme, extracted from egg whites, is an enzyme that cleaves bacterial cell walls. A 20.0-mg sample of this enzyme is dissolved in enough water to make 225 mL of solution. At 23°C the solution has an osmotic pressure of 0.118 mm Hg. Estimate the molar mass oflysozyme. 

To identify the specific aldehyde that is actually involved in the light-emitting reaction of living luminous bacteria, Shimomura et al. (1974a) extracted and purified the aldehyde from 40 g each of the bacterial cells of P. phosphoreum, Achromobacter (Vibrio or Photobacterium) fischeri, and an aldehydeless mutant of A. fischeri. The aldehyde fractions were purified, and then oxidized with Tollens reagent (silver oxide dissolved in ammonia) to convert the CHO group into the COOH group. Then the acids obtained were analyzed by mass spectrometry. The results indicated that P. phosphoreum had contained a mixture of aldehydes dodecanal (5%), tetradecanal (63%) and hexadecanal (30%), as shown in Table 2.2. Thus, tetradecanal was clearly predominant in... 

An improved method of producing recombinant aequorin was devised based on the fact that the expression of the peak amount of apoaequorin in bacterial cells occurs several hours before the secretion into culture medium (Shimomura and Inouye, 1999). The cells containing apoaequorin in the periplasmic space, before secretion, are extracted under a very mild condition and, at the same time, converted into aequorin. The purification of the extract by two steps of column chromatography yields a high-purity preparation of recombinant aequorin. 

Tamplin et. al. (54) observed that V. cholerae and A. hydrophila cell extracts contained substances with TTX-like biological activity in tissue culture assay, counteracting the lethal effect of veratridine on ouabain-treated mouse neuroblastoma cells. Concentrations of TTX-like activity ranged from 5 to 100 ng/L of culture when compared to standard TTX. The same bacterial extracts also displaced radiolabelled STX from rat brain membrane sodium channel receptors and inhibited the compound action potential of frog sciatic nerve. However, the same extracts did not show TTX-like blocking events of sodium current when applied to rat sarcolemmal sodium channels in planar lipid bilayers. 

Fractionation. The process by which components are extracted firm bacterial eells or from the medium in whieh the baeteria are grown and obtained in a purified form. The polysaccharide antigens of Neisseria meningitidis are separated from the bacterial cells by treatment with hexadecyltrimethylammonium bromide and those of Streptococcus pneumoniae with ethanol. The purity of an extracted material may be improved by resolubilization in a suitable solvent and precipitation. After purification, a component may be dried to a powder, stored indefinitely and, as required, incorporated into a vaccine in precisely weighed amounts at the blending stage. 

To check if PemB is surface exposed, E. chrysanthemi cells were subjected to proteolysis. Treatment of the cell suspension with trypsin, proteinase K or chimotrypsin at a concentration of 0.1 to 1 mg/ml for 1 h did not cause PemB proteolysis or its liberation into the medium. Cell pre-treatment with EDTA-lysozyme, which renders the periplasmic proteins accessible to proteases, gave no effect. PemB was also resistant to proteolytic digestion in extract of cells disrupted by sonication or in a French press. Only addition of Triton X-100 (up to 0.1%) causing formation of the micelles with PemB lead to a quick proteolyis of this protein (data not shown). In another approach to analyse the PemB exposition, bacterial cells were labelled with sulfo-NHS-biotin. This compound is unable to cross membranes and biotinylation... 

Allen JR, SA Ensign (1996) Carboxylation of epoxides to 3-keto acids in cell extracts of Xanthobacter strain Py2. J Bacterial 178 1469-1472. 

Brune A, B Schink (1990) Pyrogallol-to-phloroglucinal conversion and other hydroxyl-transfer reactions catalyzed by cell extracts of Pelobacter acidigallici. J Bacterial 172 1070-1076. 

Apajalahti JHA, MS Salkinoja-Salonen (1987b) Complete dechlorination of tetrachlorohydroquinone by cell extracts of pentachlorophenol-induced Rhodococcus chlorophenolicus. J Bacterial 169 5125-5130. 

PTLC was also used for the separation of lipid components in pathogenic bacteria. Mycobacterium avium has a requirement for fatty acids, which can be fulfilled by palmitic or oleic acid, and these fatty acids are then incorporated into triagylglycerols [80]. PTLC was used for the separation of fatty acids and triacylglycerols in the extracts of these bacterial cells to study the lipid classes in the bacterial cells cultured under different growth conditions. 

DNA,83 which produces one million or more copies of amplified DNA in a short time. For identification of bacteria, PCR can be used to amplify DNA either after extraction from a sample or after lysis of the cells.83,84 Methods using washing, filtration, or magnetic beads with specific antibodies can be used to collect bacterial cells for PCR.85,86 PCR can be modified for the detection of bacteria from various sources32 and can even amplify DNA from dead cells.87... 

Sample preparation used to extract proteins from cells prior to analysis is an important step that can have an effect on the accuracy and reproducibility of the results. Proteins isolated from bacterial cells will have co-extracted contaminants such as lipids, polysaccharides, and nucleic acids. In addition various organic salts, buffers, detergents, surfactants, and preservatives may have been added to aid in protein extraction or to retain enzymatic or biological activity of the proteins. The presence of these extraneous materials can significantly impede or affect the reproducibility of analysis if they are not removed prior to analysis. 

Vaidyanathan, S. Kell, D. B. Goodacre, R. Flow-injection electrospray ionization mass spectrometry of crude cell extracts for high-throughput bacterial identification. J. Am. Soc. Mass Spectrom. 2002,13,118-128. 

The method of pure polymer recovery from the biomass prior to characterization can influence the molecular weight of the polymer significantly. Extraction of PHB-bacterial cells with organic solvents yields polymers with higher molecular weight compared to sodium hypochorite treatment [44-46]. Pretreatment of the biomass with a surfactant prior to hypochlorite digestion... 

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