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Biomedical subjects

E M Gonikberg

Publications and source records attributed to E M Gonikberg.

At least 19 recordsLinked to original sources

[A novel method of transfection of marine bacteria Pseudoalteromonas espejiana with deoxyribonucleic acid of bacteriophage PM2].

DNA of bacteriophage PM2 is a convenient test object for studying DNA-damaging genotoxic agents. The extent of DNA damage can be estimated by the ability of damaged DNA for transfection of host cells, marine bacterium Pseudoalteromonas espejiana (Pae), str. BAL-31. The efficiency of transfection of Pae lines maintained for long periods without freezing was found to be very low upon the use of a widely accepted transfection method developed by van der Schans et al. (1971). Such cultures grown in a medium with 10 mM Ca2+ standard for Pae contained cell aggregates and exopolymer material. Pae was found to be capable of growing in a medium without the calcium supplement in the presence of chelator EGTA (low-calcium medium, LCM). After growth in LCM, cells did not aggregate, cultures lacked the activity of nuclease BAL, and transfection efficiency of cells grown in LCM drastically increased. Based on these results, a novel procedure of transfection with an efficiency of 2 x 10(4)-2 x 10(5) infectious centers per microgram of PM2 DNA was developed.

Corticoviridae↗

[Comparative study of cytosolic proteins in normal and neoplastic ovaries by polyacrylamide electrophoresis].

The report discusses the data on cytoplasmic protein occurrence in ovarian tissue (106) obtained by polyacrylamide gel electrophoresis. They were identified in normal ovarian tissue (15), benign tumor (16), primary malignant tumor (55) and tumor tissue after chemotherapy (20). A band was detected in the area of molecular 36 cD in malignant epithelial tumor tissue which was not seen in either normal, benign tumor tissue or after successful chemotherapy.

Adult↗

[Spontaneous mutants of Alteromonas espejiana, resistant to kanamycin and bleomycin].

An attempt was made to induce insertions in marine bacterium Alteromonas espejiana Bal-31 (Ae) using the TnphoA transposon. The Ae mutants selected on a kanamycin-containing medium after conjugation of the Ae with the transposon donor, Escherichia coli SM10(pRt291), were resistant not only to kanamycin (Kn), but also to bleomycin (Bm), and were sensitive to tetracycline. Although the mutants were phenotypically similar to insertion mutants, the mutations appeared to be spontaneous. The sensitivity of spontaneous Kmr Ae mutants selected at various Km concentrations to Bm was investigated. The mutants selected at low Km concentrations were resistant to Bm, whereas those selected at high Km concentrations were sensitive to Bm. The possible mechanisms underlying the dual resistance to Bm and aminoglycosides in bacteria are discussed.

Alteromonas↗

[Loss of adsorptive ability--the reason for inactivation of the RM2 bacteriophage during storage].

The kinetics of bacteriophage PM2 inactivation at storage was compared with the kinetics of bacteriophage adsorption on Alteromonas espejiana BAL-31 host cells. Adsorption ability and infectivity are lost with the same rate at temperatures 4-28 degrees C suggesting the loss of adsorption ability to result in bacteriophage inactivation. At higher temperatures infectivity is lost more rapidly than the ability of adsorption. The single hit kinetics of adsorption ability loss suggests the simple model of independent inactivation of 12 antireceptors located at the tops of icosaedric capsid to be erroneous. At bacteriophage inactivation the major port of protein I, a fragment of antireceptors, is preserved in the capsid composition.

Bacteriophages↗

[Inactivation of bacteriophage PM2 during storage].

The kinetics of bacteriophage inactivation in the medium that is optimal for its storage has been studied at temperatures from 4 to 55 degrees C. The plot of Arrhenius dependence of the constant of inactivation rate consists of the two linear parts with the energies of activation Ea = 25 kcal/mol for 4-37 degrees C and Ea = 91 kcal/mol for 37-55 degrees C. The DNA of inactivated bacteriophage remained mostly in superspiralized form and completely preserved its biological activity as tested by transfection in spheroplasts. The analysis of inactivation kinetics suggests ageing of virions cultivated at 4 degrees C. The addition of watersoluble antioxidant amoxipin did not change the inactivation kinetics. The addition of antioxidant ionol with twin-80 increased the inactivation that was paralleled by the bacteriophage DNA degradation.

Bacteriophages↗

[Cleavage of bacteriophage PM2 DNA by S1 nuclease].

The kinetics of cleavage of superhelical PM2 DNA by the single strand-specific S1 nuclease is studied at various salt concentrations (0.01--1 MNaCl) by electrophoresis in neutral and alkaline agarose gels. Cleavage of different DNA forms (superhelical DNA I, relaxed circular DNA II and linear DNA III) is described by the kinetic equations of the first order, and respective rate constants k1, k2, and k3 are determined at all salt concentrations used. It is shown that a high salt concentration (not lower than 0.2 MNaCl) is necessary for high specificity of S1 nuclease action on DNA regions with impaired base pairing. Examination of the S1 nuclease preparation action on superhelical PM2 DNA may be used as a convenient and reliable assay of its specificity.

DNA, Bacterial↗

DNA replication and head assembly in bacteriophage T4.

Phage DNA was accumulated in cells of E. coli B, infected with the phage T4DtsLB3 (gene 42), without the synthesis of late proteins (in the presence of chloramphenicol). Then (stage II), chloramphenicol was removed and further replication of the phage DNA suppressed with hydroxyurea and by simultaneously raising the temperature to 40 degrees. The media M9 or M9 with 1% amino acid were used; the times of addition of chloramphenicol and the hydroxyurea concentration were also varied. It was also shown that in medium M9, at stage II, chiefly early proteins were synthesized. In the medium containing amino acids, at stage II the following was observed: 1) DNA synthesis was entirely suppressed and a degradation of DNA occurred; 2) both early and late proteins were synthesized, with a predominance of the latter; 3) an assembly of the elements of the phage tails and capsids occurred without the neck and flagellum, and a small number of phage particles were also found; 4) the capsids, isolated in a sucrose density gradient after lysis with chloroform, contained the proteins Palt, P20, P23, P24, several unidentified proteins, and did not contain Pwac, P23, and P22, 5) the yield of viable phage varied from 0.05 to 15% per cell. Thus, the entire morphogenesis of T4 phage can occur without accompanying replication of phage DNA.

Chloramphenicol↗

[Photoinactivation of bacteriophage PM2 by cyanine dyes].

Photoinactivation of the lipid-containing bacteriophage PM2 by visible light and cyanine dyes (carbo- and dicarbocyanines), aluminum phtalocyanine tetrasulfonate and methylene blue was studied. It was concluded that cyanine dye aggregates adsorbed on phage particles and oxygen are essential for phage photoinactivation.

Aluminum↗

[Measuring the bleomycin-induced paired damage of two DNA chains by a method of psoralen photocross-linking].

DNA of bacteriophage PM2 was allowed to react with bleomycin in the presence of Fe(II) and oxygen and the "paired" DNA lesions of two types were measured: (1) double-strand breaks, (2) lesions converted to double-strand breaks after introducing into the DNA a large number of psoralen cross-links (about 10(-2) per base pair) and alkali treatment. The mean numbers of each lesion type per DNA molecule are found to be proportional to the square of bleomycin concentration over the range of 3 X 10(-7) to 3 X 10(-6) M. These findings indicate that paired lesions are formed as a result of action of two bleomycin molecules at the same DNA site.

Cross-Linking Reagents↗

[Kinetic parameters of superhelical DNA cleavage by endonuclease S1].

Major kinetic parameters of endonuclease S1 were determined on superhelical bacteriophage PM2 DNA and on relaxed nicked circular PM2 DNA. At 37 degrees and 0,25 M NaCl, the Michaelis constants were respectively 1.7 . 10(-8) M and 1 . 10(-9) M, and catalytic constants were respectively 0.36 sec-1 and 1.2 . 10(-2) sec-1. The inhibition of the enzyme reaction by its product was detected.

Aspergillus oryzae↗