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Biomedical subjects

J Korb

Publications and source records attributed to J Korb.

At least 19 recordsLinked to original sources
PubMed1990

Binding properties of avian retroviral proteins. I. Preparation and basic characterization of ASLV NC(p12) and MA(p19).

Using SP-Sephadex column chromatography we isolated from an avian retrovirus, AMV(MAV), nucleic acid-binding proteins ASLV NC(p12) and MA(p19). As shown by several criteria, namely SDS-PAGE, PR(p15) protease activity, and nucleic acid binding assay with the use of both ss and ds DNAs, our NC(p12) and MA(p19) isolates are virtually pure proteins mutually not cross-contaminated. Rabbit anti-NC(p12) and anti-MA(p19) sera which we prepared did not cross-react mutually. We conclude that both NC(p12) and MA(p19) and antibodies against them are adequately pure preparations for investigating their nucleic acid binding specificities towards AMV(MAV) genomic RNA and MAV-1 proviral DNA using electron microscopy supported by computer analysis of electron micrographs.

Animals↗
PubMed1990

Binding properties of avian retroviral proteins. II. Binding of protein ASLV NC(p12) to viral RNA and proviral DNA.

Binding of the major avian retroviral nucleocapsid protein ASLV NC(p12) to the MAV-1 (myeloblastosis-associated virus) proviral dsDNA and viral ssRNA was analysed using electron microscopy. Specificity of the binding was estimated by special computer programs. The NC(p12) protein bound to MAV-1 proviral dsDNA (clone pAT153--MAV-1), but specificity of this binding was not found by computer evaluation. NC(p12) also bound to nondenatured 70S viral RNA at a rate of 25 +/- 3 molecules per RNA molecule. When this RNA was denatured either before or after the complexing, it showed no binding affinity for the protein. This result implies that preserved secondary structure of the viral RNA was required for the binding.

Animals↗
PubMed1990

Binding properties of avian retroviral proteins. III. Binding of protein ASLV MA(p19) to viral RNA and proviral DNA.

The binding of the avian retroviral matrix protein ASLV MA(p19) to homologous viral ssRNA and proviral dsDNA was analysed using electron microscopic methods combined with a special computer evaluation. No binding affinity of MA(p19) to homologous nondenatured or denatured viral RNA was found. In contrast, ASLV MA(p19) was shown to have one specific binding site on MAV-1 proviral dsDNA at position 6795 +/- 345 bp from the 5' end of the molecule. A second specific binding site was found in a cellular sequence.

Animals↗
PubMed1989

[Non-imported conjunctivitis with detection of Chlamydia trachomatis].

The authors describe conjunctivitis caused by Chlamydia trachomatis. The diagnosis was established by isolation on tissue cultures with a typical finding of intracytoplasmatic inclusions. Serum antibodies were slightly elevated, the complement binding reaction was 1:16, ELISA 1:64. From the clinical aspect trachoma was not involved. The patient was not in contact with foreigners from endemic areas. The disease was cured only after general administration of Tetracycline. The authors maintain that the diagnosis and prevention of ocular Chlamydia infections in adults and neonates should be extended.

Chlamydia trachomatis↗
PubMed1983

Structural analysis of RNA subunits from avian myeloblastosis virus (AMV).

Subgenomic fragments were released from purified 35S AMV RNA using strongly denaturing conditions of 78% formamide and 3.9 M urea at 53 degrees C. The fragments were characterized by sedimentation and electron microscopic analysis. The presence of five distinct, reproducible size classes of RNA molecules with mean lengths 1.79 +/- 0.07 micron, 1.36 +/- 0.09 micron, 1.03 +/- 0.09 micron, 0.7 +/- 0.08 micron, and 0.42 +/- 0.09 micron were demonstrated. Analysis of poly(A)+ and poly(A)- RNA fragments revealed that splitting of 35S AMV RNA by strongly denaturing agents is not entirely random. Only one main distinct site of preferential splitting of the AMV RNA was located at 1.8 kb from the 3' end of the genome. The other three sites located at 3.2 kb, 4.5 kb, and 5.9 kb from the 3' end were much less distinct and were masked by random degradation of the molecules. The amount of poly(A)- RNA fragments of preferential lengths was increased by additional splitting of RNA molecules at preferential sites. No proteosynthetic activity was detected when subunits obtained by strong denaturation of 35S AMV RNA were analysed in a cell-free protein synthesis system.

Avian Leukosis Virus↗
PubMed1982

Studies on the structure of avian myeloblastosis virus (AMV) RNA. I. Factors affecting electron microscopic visualization of AMV RNA.

Factors affecting the visualization of single-stranded (ss) avian myeloblastosis virus (AMV) RNA by the basic protein film technique under strong denaturation conditions were studied. The basic parameters of the electron microscopic picture of ss RNA, involving the visibility, concentration and contour lengths of molecules, were found to be significantly dependent on the (1) quality of denaturing components used for full extension of AMV RNA molecules and (2) spreading and formation of the surface film. Under the conditions used, the presence of artificial secondary structures of AMV RNA molecules were dependent on batches of formamide and urea in the denaturing mixture. The spreading and formation of the surface film were affected by hydrophilicity of glass spreading ramps, speed of spreading, amount of spreading solution, amount of protein in the hyperphase, type of cytochrome c and the presence of substances influencing surface tension. The optimal conditions and spreading arrangement for visualization of ss AMV RNA are described.

Avian Leukosis Virus↗
PubMed1982

Studies on the structure of avian myeloblastosis virus (AMV) RNA. II. Integrity of the RNA in dependence on RNA isolation and virus propagation.

While sedimentation analysis revealed no basic changes in the sedimentation characteristics of 60-70 S avian myeloblastosis virus (AMV) RNA isolated under different conditions from two sources, electron microscopy exhibited differences in the length distributions of this RNA. Based on the length distribution profiles, phenol extraction showed a higher degradation effect in comparison with direct gradient centrifugation of viral lysates. Proteinase K appeared to be a more suitable nuclease inhibitor than diethylpyrocarbonate during the isolation of AMV RNA. Fundamental differences between length distributions of 60-70 S RNA from plasma AMV and that from AMV propagated in cultured leukaemic myeloblasts were found. Precipitation with ethanol had no effect on the length distribution of AMV RNA. Nor did storage of leukaemic chicken plasma at -70 degrees C and one thawing substantially influence the lengths of AMV RNA. The heterogeneous length distributions of 60-70 S AMV RNA molecules with preferential size-classes were independent of virus propagation and RNA isolation. The results support the idea of preferential splitting of AMV RNA at specific sites.

Avian Leukosis Virus↗
PubMed1982

Studies on the structure of avian myeloblastosis virus (AMV) RNA. III. Electron microscopic definition of secondary structure.

The secondary structure of avian myeloblastosis virus (AMV) RNA was characterized by electron microscopy under moderately denaturing spreading conditions. Under denaturation by aqueous 44% formamide or 77% formamide in the presence of salts, partly stretched RNA molecules with measurable double-stranded regions were observed. This approach allowed the localization from 5 to 11 regions of preserved secondary structure on AMV RNA molecules. Topographic analysis revealed a nonrandom occurrence of stable secondary structures in several prevalent regions. These regions with higher secondary structure stability revealed certain similarity to hairpin structures localized by electron microscopy on Rous sarcoma virus RNA or to highly structured regions found on this RNA by T1 ribonuclease oligonucleotide analysis.

Avian Leukosis Virus↗
PubMed1982

Electron microscopic studies on the structure of 60-70S RNA of avian myeloblastosis virus.

Structural properties of the 60-70S RNA complex of avian myeloblastosis virus (AMV) were analysed in electron microscope after treatment under a set of non-denaturing, gently and strongly denaturing conditions. By selected denaturing conditions, the significant fraction of 60-70S AMV RNA molecules revealed partially unfolded structures either in a dimer or a more complex form and in a length corresponding to mol. wt. of 5.6 X 10(6). The typical dimers contained a characteristic central structure connecting the subunits and similar to those described for Rous sarcoma virus (RSV) and mammalian retrovirus RNAs. This dimer linkage in the AMV genome occurred at 384 +/- 43 nucleotides from one end of each subunit. Besides partially unfolded complexes, collapsed structures and extended linear molecules were observed. The length of majority of the linear molecules had reached a half of that of the partially unfolded complexes corresponding to the mol. wt. of monomers estimated under conditions of strong denaturation to be 2.8 X 10(6). Based on our findings, we conclude that the genome of AMV shares the dimer structure with RSV and mammalian retroviruses. We also conclude that the secondary structure of AMV RNA molecule is more labile than that of RNA of mammalian retroviruses.

Avian Leukosis Virus↗
PubMed1981

Mathematical analysis of the oncornavirus maturation process (virion RNA conversion and morphological condensation).

The rate of the maturation process of avian myeloblastosis virus experimentally estimated on the basis of genomic viral RNA conversion and morphological transition of virions was mathematically analysed. Three mathematical models were suggested and fitted to experimental data. It was found that: (a) model of simple kinetics (Model 1) does not agree with experimental data. Therefore, two hypotheses were considered in further mathematical modelling: (b) virions are identical in time of budding: maturation is dependent on the presence of a virion component which is degraded with time (Model 2). This model agrees with experimental data in all stages of the maturation process. (c) Virions are released from cells at different stages of assembly (Model 3). This model differs from experimental data especially in early stages of maturation. The hypothesis used for the construction of Model 2 seems to be the most plausible to explain the maturation process and is in agreement with data of murine leukemia virus maturation which was found to be accomplished by cleavage of p70 precursor protein.

Kinetics↗
PubMed1981

Electron microscopic characterization of avian myeloblastosis virus RNA by the non-protein technique of spreading.

The quantitative analysis of the behaviour of retroviral RNA (AMV RNA) under the conditions of the non-protein technique of electron microscopic visualization on a mono-molecular film of BAC was performed. This technique resulted in visualization of intact molecules the mean length of which was 12% larger in comparison with molecules spread by the cytochrome c method. The method was found to be extremely sensitive to the surface properties of the supporting membrane which distinctly affect the shape and size of molecules. Arrangement of the surface potential of the supporting foil by means of ethidium bromide led to high reproducibility of RNA molecule stretching and to an increase in their length. The conditions were worked out in which extended linear RNA molecules were visualized, even under gentle denaturation. These conditions represent a suitable approach to the electron microscopic visualization of the protein--AMV RNA complexes.

Avian Leukosis Virus↗
PubMed1978

Size and secondary structure of avian myeloblastosis virus associated ribosomal RNA: comparison with cellular and precursor ribosomal RNA.

Ribosomal RNA isolated from ribosomes present inside avian myeloblastosis virus (AMV) was characterized by electron microscopy using the formamide-urea spreading technique. The molecular weight and the secondary structures were compared with those of r-RNA and precursor r-NA isolated from host cells, the leukemic myeloblasts. The molecular weight of viral r-RNA (1.62 +/- 0.18 X 10(6) and 0.69 +/- 0.10 X 10(6)) and the molecular weight of cellular r-RNA (1.63 +/- 0.18 X 10(6) and 0.67 +/- 0.09 X 10(6)), the latter obtained from avian myeloblasts, were found to be identical and comparable with the molecular weight of chicken liver r-RNA. Likewise, the secondary structures of viral r-RNA were identical to those of cellular r-RNA. The postulated possible precursor character of viral r-RNA was excluded, since the molecules of viral r-RNA do not show any similarity to those of precursor r-RNA. Previously observed differences in behavior of viral and cellular (myeloblastic) r-RNA in sedimentation and electrophoretic mobility are discussed.

Animals↗
PubMed1977

Electron microscopy of the segmented RNA genome ofLa Crosse virus: absence of circular molecules.

The three species of single-stranded RNA present in La Crosse virus were examined in the electron microscope. Because large amounts of contaminating cellular DNA are copurified with the virus despite extensive attempts to purify the virus, it was necessary to use procedures that eliminated the bulk of this DNA before the viral RNA was analyzed. When this was done, the modal lengths of La Crosse virus RNA were 0.4, 2.0, and 3.1 mum. These lengths correspond well to their known molecular weights of 0.4 x 106, 1.8 x 106, and 2.9 x 106. Under the denaturing conditions used to permit complete spreading of these single-stranded RNA molecules, no single-stranded circular molecules are observed. Therefore, the circular nucleocapsids present in La Crosse virus and some other bunyaviruses do not appear to be due to convalent linkage of the ends of the RNA genome.

Cell Line↗
PubMed1977

Electrophoretic analysis of the molecular weight of murine mammary tumor virus RNA.

Molecular weight determinations of native and subunit RNAs of murine mammary tumor virus (MuMTV), a type B oncornavirus, were performed by polyacrylamide gel electrophoresis and compared with molecular weights of well-characterized avian cellular RNAs and tobacco mosaic virus RNA. From extrapolations of semilog plots of the molecular weights of the standard RNAs versus relative electrophoretic mobilities and Ferguson plots, the subunit and native RNAs of MuMTV were found to possess molecular weights of 2.93 X 10(6) and 6.45 X 10(6), respectively. These data support the assumption that two subunit molecules comprise the native RNA of MuMTV.

Animals↗