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D Sagher

Publications and source records attributed to D Sagher.

31 records · Page 2Linked to original sources
PubMed1979

Effect of benzo[a]pyrene-diolepoxide on infectivity and in vitro translation of phage MS2 RNA.

Previous studies have shown that alkylation of MS2 RNA by certain derivatives of polycyclic aromatic hydrocarbons renders it noninfectious. Since phage RNA serves as a template for translation and transcription, either of these RNA-directed processes, or both, could be responsible in vivo for the inhibition of phage replication by metabolically activated hydrocarbons. The present study correlates the degree of inhibition of MS2 RNA infectivity, at various levels of alkylation by (+/-)-trans, 7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzol[a]pyrene, with the translation efficiency in vitro of the same alkylated RNA for the synthesis of viral synthetase and of maturation and coat proteins. The results indicate that dihydroxyepoxy-tetrahydrobenzo[a]pyrene modification of MS2 RNA impairs its template capacity for the synthesis of phage-specific proteins; this inhibition is insufficient, however, to account for the loss of RNA infectivity at lower molar ratios of alkylation. For the three viral proteins synthesized in vitro, the translation of RNA synthetase is much more sensitive to MS2 RNA modification than either coat or maturation protein synthesis. Our results also indicate that the loss of viral RNA infectivity follows a single-hit inactivation mechanism, whereas several alkylation events in the viral RNA synthetase cistron may be necessary to block translation of this gene product.

Alkylation↗
PubMed1976

Large subunit ribulosebisphosphate carboxylase messenger RNA from Euglena chloroplasts.

RNA from wild-type Euglena, aplastidic mutant cells, and purified chloroplasts was chromatographed on oligo(dT)-cellulose at 4 degrees. The poly(A)-containing and poly(A)-lacking fractions from each were then tested in a cell-free protein-synthesizing system from wheat germ for their abilities to specifically stimulate synthesis of large subunit ribulosebisphosphate carboxylase [EC 4.1.1.39; 3-phospho-D-glycerate carboxy-lyase (dimerizing)]. The large subunit polypeptide (59,000 molecular weight) was identified in the in vitro reaction by two-dimensional electrophoresis involving isoelectric focusing and size filtration on polyacrylamide gels. Template activity for the large subunit was detected in wild-type cells but not in aplastidic mutant strains; it was highly enriched in the isolated chloroplasts. This messenger was present only in the poly(A)-lacking RNA fraction, where it constituted the most prominent template species of chloroplast RNA. The large subunit message was freed of considerable non-messenger RNA contamination and localized to the 10-20S fraction by sucrose gradient centrifugation.

Carboxy-Lyases↗
PubMed1975

Ribulose Diphosphate Carboxylase from Autotrophic Euglena gracilis.

Ribulose 1,5-diphosphate carboxylase (RUDPcase) from autotrophically grown Euglena gracilis was purified to homogeneity as measured by analytical ultracentrifugation, polyacrylamide gel electrophoresis, and immunoprecipitation reactions. The enzyme represented about 9% of total protein and 24% of soluble protein in the autotrophic cell. Light-grown, heterotrophic cells seemed to contain considerably less RUDPcase. Native carboxylase from autotrophic Euglena showed an s(20, w) at low protein concentrations of 17 to 17.5, suggesting a molecular weight of >500,000 daltons. Upon denaturation, the enzyme dissociated into two subunits having different amino acid compositions and molecular weights of 59,000 and 12,000 daltons. Based upon the amino acid mass ratios, a quaternary organization of 7 to 8 large and 8 to 10 small subunits per native enzyme molecule was indicated.The phylogenetic relationship of carboxylase from Euglena and from three higher plants was investigated. In general, the size, subunit formation, and quaternary structure of RUDPcase from the various sources seemed to be similar. A partial immunochemical reaction between anti-RUDPcase serum from Euglena and the enzymes from lettuce, cucumber, and New Zealand spinach suggested that the algal and higher plant carboxylases were related but not identical. This was borne out by amino acid analyses which showed a close correspondence between the large, but not the small, subunits of Euglena and lettuce.

Journal Article↗
PubMed1974

Poly(A)-associated RNA in plants.

The RNA associated with poly(A) sequences from Euglena gracilis and Vicia faba has been isolated by binding to millipore filters and characterized by sedimentation velocity centrifugation and electrophoretic mobility. Poly(A)-associated RNA as isolated in solution was highly aggregated. When denatured, it sedimented as a broad peak with a mean value of 16-18 S. This RNA was shown to be covalently linked to poly(A) sequences which are 150-250 nucleotides long. Our size estimates for plant poly(A) and poly(A)-associated RNA are similar to those obtained for animal cells.

Animals↗
PubMedDate not supplied

The role of DNA polymerase in base substitution mutagenesis on non-instructional templates.

In vitro DNA synthesis on phi X174 or M13 templates with non-instructional lesions such as UV dimers or AP (apurinic/apyrimidinic) sites terminates one base before the site of the lesion when synthesis is catalyzed by T4 DNA polymerase or E. coli polymerase I. E. Coli polymerase I also produces termination bands at the site of AP lesions. Substitution of Mn2+ for Mg2+ and increasing the concentration of dNTP's results in elongation of the newly synthesized strand opposite the site of the lesion and beyond. Purine deoxynucleoside triphosphates are utilized for insertion opposite lesions to a greater extent than are pyrimidine deoxynucleoside triphosphates. Deoxy ATP is used almost exclusively for elongation opposite AP sites with pol I-Klenow fragment in the presence of Mg2+. We suppose that these results illustrate the previously observed greater affinity of polymerases under template-free conditions for purine nucleotides. We also suppose that the results can be used to account for mutagenic base selection on noninstructional DNA templates. If purines are preferentially selected by polymerases, then treatments which inactivate pyrimidines will lead to an excess of transitions whereas inactivation of purines will produce more transversions. Data in the literature support this hypothesis.

Bacteriophage phi X 174↗