Abstract: A method is provided for making synthetic capped RNAs. These compounds serve as substrates for the virally encoded endonuclease associated with influenza virus. We are able to assay for this unique and specific viral activity of cleavage of a capped RNA in vitro. Therefore, screening of inhibitors of this activity is possible. In addition, short non-extendible (due to their length or because of the modification of the 3′-end of the oligo, i.e. 3′-dA) RNAs are potent inhibitors of the cleavage of capped RNAs by influenza endonuclease. Finally, these compounds may be used to investigate viral and cellular mechanisms of transcription/translation or mRNA maturation.

Abstract: A method is provided for making synthetic capped RNAs. These compounds serve as substrates for the virally encoded endonuclease associated with influenza virus. We are able to assay for this unique and specific viral activity of cleavage of a capped RNA in vitro. Therefore, screening of inhibitors of this activity is possible. In addition, short non-extendible (due to their length or because of the modification of the 3'-end of the oligo, i.e. 3'-dA) RNAs are potent inhibitors of the cleavage of capped RNAs by influenza endonuclease. Finally, these compounds may be used to investigate viral and cellular mechanisms of transcription/translation or mRNA maturation.

Abstract: The present invention provides rapid accurate sensitive assays specific for the detection of at least one a single stranded oligonucleotide produced by the action of an enzyme on a substrate. The assays are useful to detect the presence in a sample of an enzyme which acts on an oligonucleotide substrate to generate a single stranded oligonucleotide product and to detect inhibitors of such an enzyme.

Abstract: At low temperature, the protease of human cytomegalovirus (HCMV) is rendered highly active as a dimer of identical units. The protease is useful as a screening tool for HCMV antivirals as well as a diagnostic tool for diseases resulting from HCMV infection.

Abstract: A method is provided for making synthetic capped RNAs. These compounds serve as substrates for the virally encoded endonuclease associated with influenza virus. We are able to assay for this unique and specific viral activity of cleavage of a capped RNA in vitro. Therefore, screening of inhibitors of this activity is possible. In addition, short non-extendible (due to their length or because of the modification of the 3'-end of the oligo, i.e. 3'-dA) RNAs are potent inhibitors of the cleavage of capped RNAs by influenza endonuclease. Finally, these compounds may be used to investigate viral and cellular mechanisms of transcription/translation or mRNA maturation.

Abstract: An assay for the influenza virus endonuclease has been developed which involves DNA polymerase-catalyzed extension of the viral endonuclease cleavage product using labeled nucleotides and a DNA template containing a 3' region complementary to the product joined to a 5' region consisting of repeated residues. The DNA polymerase coupled assay does not involve gel electrophoretic separation and is amenable to high volume screening of potential inhibitors. Another key feature of the assay is that it is sensitive enough to detect 200 attomoles of product.

Abstract: Native Herpes Simplex Virus type 1 (HSV type 1) protease has a rather low activity. It has been found however, that the presence of a kosmotropic anion can increase the activity of the protease over 10-fold. This has many advantages, as now assays for protease activity may be run in very small (micromolar and less) amounts of reagents and substrates, and the assays can be run quickly. Activate HSV-1 protease represents a conformational change from native protease.

Abstract: A method of purifying proteases from the Herpes Simplex Virus 1 and 2 is taught. A precursor protease is cloned in a host cell and then separated from the cell culture using cation exchange chromatography. The resulting protease is subjected to autoprocessing conditions and an autolytic cleavage occurs, resulting in mature protease.