title: RNase H: specificity, mechanisms of action, and antiviral target.
abstract: The Ribonuclease (RNase) H is one of the four enzymes encoded by all retroviruses, including HIV. Its main activity is the hydrolysis of the RNA moiety in RNA-DNA hybrids. The RNase H ribonuclease is essential in the retroviral life cycle, since it generates and removes primers needed by the Reverse Transcriptase (RT) for initiation of DNA synthesis. Retroviruses lacking RNase H activity are noninfectious. Despite its importance, RNase H is the only enzyme of HIV not yet targeted by antiretroviral therapy. Here, we describe functions and mechanisms of RNase H during the HIV life cycle and describe a cleavage assay, which is suitable to determine RNase H activity in samples of various kinds. In this assay, an artificial, fluorescence-labeled RNA-DNA hybrid is cleaved in vitro by an RT/RNase H enzyme. Cleavage products are analyzed by denaturing polyacrylamide gel electrophoresis (PAGE). This assay may be used to detect the RNase H, assess the effect of inhibitors, or even activators, of the RNase H, as we have described, as candidates for novel antiretroviral agents.
authors.affiliation: Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland.
authors.forename: Karin
authors.initials: K
authors.lastname: Moelling
authors.forename: Felix
authors.initials: F
authors.lastname: Broecker
authors.forename: John E
authors.initials: JE
authors.lastname: Kerrigan
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology*; D057075:Enzyme Assays / Q000379:methods*; D015497:HIV-1 / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D008958:Models, Molecular; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D059748:Proteolysis; D054309:Ribonuclease H, Human Immunodeficiency Virus / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D019380:Anti-HIV Agents; D054309:Ribonuclease H, Human Immunodeficiency Virus