Tularik Limited

Michael Gregory Peterson

Mohanram Sivaraja

Anthony Simon Lynch

Berta Strulovici

Binoj Joseph Matthew

Vijay R Baichwal

Thomas Henkel

Heidi Giordano

High-throughput in vitro assays are provided for identifying modulators of peptidyl transferase. New solid-phase assays, related compositions, apparatus and integrated systems are provided.

High-throughput in vitro assays for modulators of peptidyl transferase

The invention provides screening assays for identifying an inhibitor of a pathogenic helicase activity, e.g. a helicase derived from a pathogenic infectious organism such as a bacterium, protozoan or fungus. Helicase activity is detected by the solid-phase, preferential capture of retained (non-liberated) single-stranded nucleic acid comprising a detectable label.

High-throughput screening assay for inhibitors of nucleic acid helicases

The invention provides methods and compositions for identifying pharmacological agents useful in the diagnosis or treatment of disease associated with the expression of a gene modulated by a transcription complex containing at least CBF1 and a transcription factor. The methods involve combining CBF1, the Epstein-Barr virus transcription factor EBNA2 or cellular homolog thereof, and a candidate pharmacological agent. This mixture is incubated under conditions whereby, but for the presence of the candidate pharmacological agent, CBF1 binds EBNA2. The absence of selective binding indicates that the candidate pharmacological agent is capable of selectively modulating the expression of a gene dependent on CBF1-transcription factor binding. The mixture may further comprise a CBF1 binding element including the nucleotide sequence: C-G-T-G-G-G-A-A. This mixture is incubated under conditions whereby, but for the presence of the candidate pharmacological agent, a transcription complex comprising both polypeptides bound, directly or indirectly to the nucleic acid is formed. The methods are particularly suited to high-throughput screening where one or more steps are performed by a computer controlled electromechanical robot comprising an axial rotatable arm and said solid substrate is a portion of a well of a microtiter plate.

Epstein-barr virus transcription factor binding assay

Pharmacological agents useful in the diagnosis or treatment of disease associated with the expression of a gene are identified in high throughput drug screening assays. The methods involve combining a labeled transcription factor, a nucleic acid coupled to a ligand, a candidate pharmacological agent and a receptor immobilized on a solid substrate, such as a microtiter plate, filter, or bead. The nucleic acid has at least that portion of a nucleotide sequence naturally involved in the regulation of the transcription of the gene which is necessary for sequence-specific interaction with the transcription factor. The resultant combination is incubated under conditions whereby the receptor is bound to the ligand and, but for the presence of said candidate pharmacological agent, the transcription factor is sequence-specifically bound to the nucleic acid. Unbound transcription factor is then removed or washed from the solid substrate and labelled, sequence-specifically bound transcription factor is detected. Incubates which include candidate agents which alter transcription factor binding deviate from control incubates in terms of label signal--typically, binding is disrupted and the signal is diminished. In a preferred embodiment, the entire process is performed by a computer-controllable electromechanical robot with an axial rotatable arm.

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Millbrae, CA, United States of America

Michael Gregory Peterson