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Biomedical subjects

D E Georgopoulos

Publications and source records attributed to D E Georgopoulos.

8 recordsLinked to original sources
PubMed2000

Use of high-performance liquid chromatographic fractionation of large RNA molecules in the assay of group I intron ribozyme activity.

Ion-pair reversed-phase high-performance liquid chromatography (HPLC), which has previously been used to fractionate double-stranded DNA molecules, can be applied to single-stranded RNA molecules in the size range of 200-1000 nucleotides. This procedure permits RNA molecules to be separated and recovered rapidly in liquid medium, thereby facilitating recovery. We have used this system to separate an in vitro transcription product containing a group I intron ribozyme from the intermediates and products of the splicing reaction, permitting rapid assay of ribozyme activity without the use of radioactivity.

Chemical Fractionation↗
PubMed2000

The [KIL-d] element specifically regulates viral gene expression in yeast.

The cytoplasmically inherited [KIL-d] element epigenetically regulates killer virus gene expression in Saccharomyces cerevisiae. [KIL-d] results in variegated defects in expression of the M double-stranded RNA viral segment in haploid cells that are "healed" in diploids. We report that the [KIL-d] element is spontaneously lost with a frequency of 10(-4)-10(-5) and reappears with variegated phenotypic expression with a frequency of > or =10(-3). This high rate of loss and higher rate of reappearance is unlike any known nucleic acid replicon but resembles the behavior of yeast prions. However, [KIL-d] is distinct from the known yeast prions in its relative guanidinium hydrochloride incurability and independence of Hsp104 protein for its maintenance. Despite its transmissibility by successive cytoplasmic transfers, multiple cytoplasmic nucleic acids have been proven not to carry the [KIL-d] trait. [KIL-d] epigenetically regulates the expression of the M double-stranded RNA satellite virus genome, but fails to alter the expression of M cDNA. This specificity remained even after a cycle of mating and meiosis. Due to its unique genetic properties and viral RNA specificity, [KIL-d] represents a new type of genetic element that interacts with a viral RNA genome.

Base Sequence↗
PubMed1996

Hybridization properties of oligodeoxynucleotide pairs bridged by polyarginine peptides.

The hybridization properties of a series of probes, based on two 9mer oligodeoxynucleotides (designated as I and II) having an appended oligoarginine chain (Rn) to produce peptide-oligonucleotide conjugates or peptide-bridged oligonucleotide pairs (e.g. Rn-I or II-Rn-I), were investigated. For the double-linked probes, we found that the peptide bridge induces the two 9mers to bind complementary single-stranded DNA or RNA targets with substantially enhanced thermal stability. The resulting hybrid with complementary DNA was found to assume a 1:1 complex in the B conformation as judged by UV mixing curves and CD spectroscopy. Complexes of single or double-linked probes with complementary RNA exhibited sensitivity to RNase H digestion. The influence of the identity and chirality of the repeating unit in the bridge, the length of the bridge, the gap size and the salt concentration on the hybridization properties of this new class of oligonucleotide probes was also studied. Our data reveal that these compounds exhibit properties that should prove useful in the development of antisense strategies.

Amino Acid Sequence↗
PubMed1993

Oligonucleotide-poly-L-ornithine conjugates: binding to complementary DNA and RNA.

On the basis of the reported enhanced antisense activity of polylysine-oligonucleotide conjugates, a synthetic 12-mer oligodeoxyribonucleotide has been coupled at its 5' terminus to a series of positively charged (delta-ornithine)n cysteine peptides. Binding between the nucleic acid-peptide conjugate and its complementary DNA target sequence was detected by the impact of complexation on the melting temperature (Tm). It was found that the Tm for the nucleic acid-peptide gradually increased with increasing net charge on the conjugated peptide. Site-directed cleavage with RNase H demonstrates that the peptide-modified oligomer also hybridizes with its RNA target sequence. Increased affinity for target mRNA with net charge was shown by a cell-free translation arrest assay.

Base Sequence↗
PubMed1987

Nucleotide phosphotransferase, nucleotide kinase and inorganic pyrophosphatase activities of killer virions of yeast.

The intracellular killer virions of yeast co-purify with an RNA polymerase activity which catalyzes the synthesis of full-length transcripts of the two viral genomic double-stranded RNA segments. This polymerase utilizes ribonucleoside diphosphates or triphosphates as substrates. The virions have other associated nucleotide-metabolizing enzyme activities, including nucleoside diphosphate kinase, adenosine monophosphate kinase, and nucleoside triphosphate phosphotransferase, an activity which catalyzes the exchange of gamma-phosphate from any ribonucleoside triphosphate with any ribonucleoside or deoxyribonucleoside triphosphate. The purified virions also contain an inorganic pyrophosphatase activity. These enzymes may allow the virus to utilize nucleotide pools distinct from those utilized in host cell transcription.

Adenosine Diphosphate↗
PubMed1986

Sequence of the M1-2 region of killer virus double-stranded RNA.

A full-length complementary DNA (cDNA) copy of the M1-2 region of the double-stranded genome of the yeast killer virus was synthesized by reverse transcription, utilizing the m in vitro transcript as template and synthetic primers for both strands. The sequence lacks any long open reading frames (ORFs). The internal portion of the M1-2 region includes the sequence that is linked to the subterminal 229 bases of the M1-1 homologous region in the S3 defective-interfering mutant of killer virus double-stranded RNA (dsRNA). Thus, the probable site at which the deletion occurred in S3 has been identified.

Base Sequence↗
PubMed1982

Arylsulfatase A activity and electrophoretic banding patterns from isolated human blood fractions.

Arylsulfatase A (ASA) activity was studied from five blood fractions, leukocytes plus platelets (LKPL), leukocytes (LK), neutrophils (N), lymphocytes (LYM), and platelets (PL). No significant difference was found among the mean ASA specific activity values for the fractions. Electrophoretic examination revealed four distinct activity bands for the LKPL and PL fractions, while the LK, N, and LYM fractions showed only a single, broad peak, which indicates that the four-band pattern is associated with enzyme obtained from platelets. The PL fraction gave a clearer, more distinct banding pattern than the other four fractions. This clearly resolved electrophoretic banding pattern of ASA from pure platelet preparations was demonstrated for variant human ASA as well. These findings may be significant to future work investigating the biochemical genetics of ASA.

Arylsulfatases↗