Immunoelectrophoresis analysis of the antigenic composition of E. coli K12 hybrids which acquired the serological specificity of S. newcastle and S. flexneri

Genetic and immunoelectrophoretic studies confirm earlier data on the presence of 2 specific antigens of acidic nature in S. newcastle; one of them is a specific thermolabile K-antigen responsible for type IV specificity of these bacteria. The data concerning the differences in the genetic determinants controlling the synthesis of O- and K-antigens in S. newcastle have been obtained. S. newcastle O- and K-antigens did not react with S. flexneri in the group serum system 3, 4, which indicates that S. newcastle are serologically isolated and form a separate taxonomic group of dysentery bacteria. The existence of cross reactions between S. flexneri and S. newcastle due to the presence of neutral R-core antigens common to these 2 species has been shown . Immunoelectrophoresis in agar is the most promising and informative method in genetic and chemical studies of the antigenic structure of bacteria.     

The decrement in light sensitivity of the isolated frog retinal rod in the presence of a phosphorylation-resistant GDP analogue of guanosine-5′-O-(2-thiodiphosphate) as a confirmation of the hypothesis about transducin activation via the transphosphorylation mechanism

The decrement in light sensitivity of the isolated frog retinal rod cell was demonstrated after a short-time perfusion with guanosine-5′-O-(2-thiodiphosphate), which is an analog of GDP that is resistant to phosphorylation by nucleoside diphosphate kinase. This decrement can be explained by the hypothesis that transducin, which is the main GTP-binding protein of the retinal rod photoreceptor of vertebrates, is activated by phosphorylation of bound GDP to GTP; this is induced by the activated rhodopsin receptor. The results can be considered as a confirmation of the proposed hypothesis.     

A Ca2+-Binding Protein that Regulates the Lifetime of the Transducin Active State

Biophysics - Abstract—It has been previously shown in the study of the kinetic behavior of cGMP-specific phosphodiesterase in preparations of the outer segments of bovine retinal rods, which...     

Brightness of yellow fluorescent protein from coral (zFP538) depends on aggregation

The yellow fluorescent protein from coral (zFP538) forms aggregates in water solutions. According to dynamic light scattering and gel filtration data, the aggregation number is approximately 1000-10000 at pH 8-9 and protein concentration 1 mg/mL. Gel filtration demonstrated that dissociation of the aggregates takes place upon dilution, and the molecular weight of the aggregates decreases with pH. Atomic force microscopy (AFM) and near-field scanning optical microscopy (NSOM) were used to obtain images of zFP538 in the solid state. It was shown that protein films are comprised of fluorescent ellipsoidal granules with a 50-300 nm major axis and a 30-130 nm minor axis. The dependence of zFP538 fluorescence on protein concentration between 1.2 x 10(-)(9) and 5.5 x 10(-)(7) M can be divided in two linear regions with different slopes indicating the existence of at least two different forms of zFP538. The fluorescence of zFP538 decreases with time upon acidification, and the decrease depends on pH and protein concentration. Between pH 3.5 and pH 5.5, relative residual fluorescence is higher for concentrated zFP538 solutions (about 10(-)(6) M) as compared with diluted ones (10(-)(7) M and below). Aggregation makes zFP538 more stable against fluorescence quenching upon acidification: the decrease in zFP538 fluorescence at protein concentration 1 mg/mL is completely reversible, unlike that observed for less concentrated solutions. This phenomenon may be due to the decrease in the freedom of chromophore mobility in zFP538 aggregates.     

Importance of electroroentengography and fluorography in the detection of bronchiectatic disease

During mass screening of population 54 patients were detected with suspected bronchiectatic disease. Photoroentgenography and electroroentgenography were performed in all of them, in 36 of them simultaneously with subsequent bronchography. On electroroentgenograms cellular deformity in the decreased lung, the presence of cellular and linear lucidity of bronchiectasia against a background of a gas-bubble of the stomach were referred to as signs of bronchiectatic disease. Signs of inflammatory exacerbation and symptoms of lung emphysema were revealed on photoroentgenograms as an addition to electroroentgenographic findings. Combined electrophotoroentgenography with subsequent bronchographic verification of the spread of changes is recommended in suspected bronchiectatic disease.     

Immunoelectrophoretic analysis of the antigenic composition of Yersinia

The antigenic composition of 24. Y. pseudotuberculosis newly isolated and reference strains, 7 Y. enterocolitica strains, as well as Y. pestis vaccine strain EV, has been studied by the method of immunoelectrophoresis in agar. The antigenic composition of these bacteria has been found to be complicated and to comprise not less than 8-11 antigens, and among them nonspecific protein antigens common for enterobacteria, the common generic antigen, the antigen common with Y. pestis, as well as O-antigens specific for each serovar are identified. Immunoelectrophoretic study has shown the possibility of Y. pseudotuberculosis O-antigen, serovar I, with Salmonella sera, serogroup A, and Y. enterocolitica 09 with brucellar and cholera sera.     

Preparation and characterization of calibration beads for sorting cells expressing a beta-lactamase gene reporter.

BACKGROUND: Modern drug discovery has been based on high-throughput screening using whole-cell assays. A prominent role has been assigned to the reporter gene technology based on a beta-lactamase and the fluorogenic substrate CCF2. Successful application of this technology requires fluorescence-activated cell sorting. We describe the preparation and characterization of calibration beads for sorting cells expressing the beta-lactamase gene using the CCF2 substrate. METHODS: To model Forster resonance energy transfer (FRET) between the coumarin donor and the fluorescein acceptor of the CCF2 reporting dye, we used activated polystyrene beads with primary amino groups. Donor and acceptor fluorophores were attached to the beads at different ratios via succinimidyl esters. The beads were characterized with a fluorescence plate reader and a flow cytometer. RESULTS: We prepared polystyrene beads with five different ratios of donor and acceptor fluorophores and beads that carried a donor or a receptor fluorophore alone. Fluorescence measurements demonstrated that the prepared beads well represent the FRET of CCF2 substrate. CONCLUSION: We have demonstrated that the prepared beads can be successfully used for the setup of fluorescence-activated cell sorting to sort cells with CCF2 reporter substrate and the beta-lactamase reporter gene.     

Kinetics of rearrangement of solvation sheath of an excited molecule of the fluorescent probe 4"-dimethylaminochalcone

Processes accompanying the quenching of the fluorescent probe 4"-dimethylaminochalcone by hydroxyl groups of the proton-donor solvent 1-butanol have been studied. The kinetics of the deactivation of the excited state of 4"-dimethylaminochalcone has been monitored from the transition absorption spectra at a time resolution of 50 fs and fluorescence decay at a time resolution of 30 ps. The data obtained allow thinking that the next picture occurs in 1-butanol. At first stage, the 4"-dimethylaminochalcone molecule in its ground state forms a hydrogen bond with an alcohol molecule. At the second stage, the absorption of light quantum and corresponding rise of the dipole moment of 4"-dimethylaminochalcone take place, the initially existing hydrogen bond is retained. The third stage consists in the rearrangement of the 4"-dimethylaminochalcone solvation shell formed by alcohol dipole molecules due to an increase of the dipole of moment 4"-dimethylaminochalcone; this rearrangement takes an energy of about 24 kJ/mol, the arrangement time constant is close to 40 ps; the initial hydrogen bond is retained. The fourth stage involves processes that lead to fluorescence quenching; their time constant is about 200 ps. Taking into account that the quenching is a much slower process than the relaxation of the solvation shell, it was supposed that the quenching is not a direct consequence of the solvation shell relaxation or the existence of the hydrogen bond formed prior to excitation. Then the fluorescence quenching of 4"-dimethylaminochalcone can be accomplished through some other processes that are observed in other fluorescent molecules: (a) rearrangement of the initial hydrogen bond from a conformation that cannot quench the fluorescence of 4"-dimethylaminochalcone to a more "effective" conformation, (b) charge transfer between the excited of molecule 4"-dimethylaminochalcone and alcohol, or (c) solvent-induced twist of the 4"-dimethylaminochalcone amino group (its withdrawal from the molecule plane) by the action of the solvent.