High-throughput screening assay for inhibitors of nucleic acid p

Details High-throughput screening assay for inhibitors of nucleic acid p High-throughput screening assay for inhibitors of nucleic acid p

1994-12-02

1997-06-03

Jones, W. Gary

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

435 5, 435 912, 536 243, 623 6, 422101, 210633, C12Q 168, C12Q 170, C12P 1934, C07H 2104

Patent

active

056353499

ABSTRACT:
The invention provides promoter specific and promoter non-specific screening assays for identifying an inhibitor of a pathogenic nucleic acid polymerase activity, e.g. an RNA polymerase derived from a pathogenic infectious organism such as a bacterium, protozoan or fungus. Generally, the methods involve forming a mixture of nucleotides, a polynucleotide template, a pathogenic polymerase candidate inhibitor of polymerase activity, where at least one of the nucleotides comprises a detectable label. The mixture is incubated under conditions whereby, but for the presence of the candidate inhibitor, the polymerase transcribes the polynucleotide template by catalyzing the polymerization of the nucleoside triphosphates into a polynucleotide having a nucleotide sequence complementary to that of the polynucleotide template. The nascent transcript is captured by a polynucleotide-selective agent immobilized on a solid substrate for subsequent washing and label detection.

REFERENCES:
Flanagan et al., "Simple Derivation of TFIID-dependent RNA polymerase II Transcription Systems from Schizosaccharomyces pombe and Other Organisms, and Factors Required for Transcriptional Activation", Proc. Natl. Acad. Sci. USA 89:7659-7663 (1992).
Ruet et al., "A Specific Assay for Yeast RNA polymerases in Crude Cell Extracts", Eur. J. Biochem. 90:325-330 (1978).
Sorger et al., "Identification and Purification of Sequence-specific DNA-binding Proteins", Protein Function--A Practical Approach Creighton, Ed. IRL Press, Oxford University Press, Oxford, NY (1989).
Stunnenberg et al., "RNA Polymerase from the Fungus Aspergillus nidulans", Eur. J. Biochem. 98:107-119 (1979).

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