Biomedical subjects
Publications and source records attributed to J Doskocil.
Recombinant baculoviruses expressing the v-myb and c-myb genes in infected insect cells were constructed. The electrophoretic mobilities of their immunoreactive products were the same as those of the authentic Myb proteins from chicken cells. The system provides a convenient source of relatively large amounts of v-Myb or c-Myb for in vitro binding studies.
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Restriction fragment patterns of G+C-rich satellites of sheep and goat DNA were compared. The 1,712 g/cm3 satellites of both species appear homologous, consisting of repeats 760 base pairs long and showing coincidence of position of primary+ EcoRI, BamHI and most BspRI restriction target sites. The EcoRI and BamHI endonucleases produce mostly monomers of the repeating unit, while oligomers prevail in the A1uI and Bg1II digests. Species-specific differences in the frequency, position and mode of distribution of secondary+ restriction target sites for EcoRI, Bg1II and A1uI were observed. Unlike the 1,712 g/cm3 satellites, the 1,723 g/cm3 component of sheep DNA and the 1,719 g/cm3 material from goat DNA appear species--specific, since no homologous material could ever be detected in the DNA of the other species.
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Prolonged repeated analyses of sera from 98 patients with systemic lupus erythematosus (SLE) and 76 patients with similar diffuse diseases of connective tissue by a radioisotope membrane binding method for the assessment of antibodies against double-stranded ribonucleic acid (anti-ds-RNA) revealed that these antibodies are most frequently encountered in patients with the evolutive form of SLE (53%), but also in approximately one fifth of patients with non- evolutive SLE and in patients with similar diffuse diseases of connective tissue. Anti-ds-RNA are not linked with stages of clinical activity; their occurrence is on the whole independent on the occurence of antibodies against ds-DNA and nuclear ribonucleoprotein. There is a very close association with the incidence of antibodies against denatured DNA and a substantially small association with antibodies against acid nuclear antigen (Sm). The occurrence of anti-ds-RNA is a favourable prognostic sign, as in the majority of patients who had such antibodies at any time in their serum, later the disease can be suppressed by treatment, while patients who never had anti-ds-RNA in serum remain for a long time in the active stage of the disease.
The administration of sonicated fractions of f2 phage polyribonucleotides caused an increased weight loss and deterioration of the clinical state in female NZB mice. Discontinuance of the treatment resulted in an improvement of both the clinical state and the genetically determined autoimmune disorders of these mice. Some potential explanations of this effect are discussed.
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Double stranded RNA obtained from non-permissive E. coli cells infected with f2-phage was tested in 5 hospitals in viral dermtoses such as herpes simplex recidivans, herpes zoster, male genikeratonconjunctivitis herpetica and conjunctivitis lignosa. The results of clinical tests indicate that the preparation of phage double stranded RNA applied topically is harmless for man and has, in the majority of cases, a beneficial effect in the disease. This conclusion was based on the judgment of physicians, opinion of patients expressed in a questionnaire, and, results of a double-blind experiment.
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The dynamics of interferon formation by an established cell line of mouse fibroblast (L cells) and by mouse peritoneal leukocytes induced by double-stranded RNA extracted from E. coli f2 phage is described. The L cells produced interferon at a lower rate, the maximum values were obtained at 12 to 20 hours after induction, and the production was ultimately dependent on the established cell line used and on the presence of DEAE-dextran during induction. The mouse peritoneal leukocytes (MPL), on the other hand, did not require DEAE-dextran and the maximum of interferon production was reached between 6 and 12 hours after induction. Both the L cell- and the MPL-interferons were purified and concentrated so that the final specific biologic activity was 100-to 300-fold higher than that of the initial preparations (1 to 5 X 10(6) interferon units per mg protein). Polyacrylamide gel electrophoresis showed similar migration profiles for the preparations of both interferons. The smaller part of the activity was situated in a broader, slow-moving peak and the greater part formed a sharp, high and fast-moving peak. Using 3H uridine-labelled f2 ds-RNA for induction of interferon it was found that one of the radioactivity zones coincided with the fast-moving activity peak of the purified and concentrated interferon.
(1) The inactivation of various forms of nucleoside transport with reagents blocking thiol groups was studied in whole cells of E. coli B. No positive correlation between the efficiency of active transport and the extent or rate of inactivation could be demonstrated. (2) The most efficient constitutive nucleoside-transporting system was found to comprise a specific thiol component characterized by low rate of inactivation with N-ethylmaleimide; the less efficient inducible transport and the facilitated diffusion of guanosine require the integrity of another thiol component which is rapidly inactivated with N-ethylmaleimide. (3) The constitutive nucleoside-transporting system is completely inactivated with T4 phage, while other modes of nucleoside transport are much less affected. (4) Inactivation of constitutive transporting system in cells exposed to N-ethylmaleimide for a limited period of time continues long after the inhibitor has been removed, indicating storage of the inhibitor in some cellular compartment. Addition of dithiothreitol stops the inactivation immediately.
Interaction of histone H 1 and models simulating histone chains was followed by monitoring the melting curves of supernatants after the sedimentation of aggregated complexes. In a mixture of two DNAs the histones reacted selectively with (A+T)-rich and non-methylated DNA, respectively. H 1 and (Ala-Lys-Pro)n also interacted preferentially with DNA in a mixture with double stranded RNA whereas (Lys30,Ala70)n did not show any selectivity. (G+C)-rich DNA in complexes showed CD spectra the intensity of which decreased with increasing DNA methylation to values comparable with these of complexes of (A+T)-rich DNA. In complexed with double stranded RNA only the polymer (Lys30,Ala70) displayed CD pattern similar to spectra of complexes with DNA. It was concluded that formation and structure of complexes depend selectively on the DNA conformation and base composition.
Molecules of single-stranded ribosomal RNA and double-stranded replicative form of phage f2 RNA (dsRNA) adopt a compact form in solutions, containing sufficiently high concentrations of salt (NaCl) and polymer (PEG). However, only in the cases of native dsRNA molecules the compact particles are characterized by a regular internal structure, which accounts for the appearance of an intense positive band in CD spectra. Heating or acidification of PEG-containing solutions of dsRNA leads to the disappearance of the intense positive CD band, which results from the "destruction" of the regular internal structure of compact particles. Comparison of properties of DNA and dsRNA compact particles formed in PEG-containing water-salt solutions suggests the existence of similar mechanisms of compactization of double-stranded polynucleotides.
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