Inhibition of the self-cleavage reaction of the human hepatitis delta virus ribozyme by antibiotics.
Human hepatitis delta virus (HDV) poses a health threat in populations where chronic hepatitis B is endemic. It is a single-stranded RNA virus of 1700 nucleotides and both genomic and antigenomic sequences contain ribozymes which are important for viral replication. Using ribozyme constructs we show that several classes of antibiotics inhibit the self-cleavage reaction of the HDV ribozyme. Antibiotics of the aminoglycoside, peptide and tetracycline classes all inhibit HDV cleavage in vitro at micromolar concentrations. Neomycin (an aminoglycoside) inhibits HDV self-cleavage with a Ki value of 28 (+/- 10) microM. Neomycin inhibition can be reversed by increasing magnesium ion concentration in a competitive manner. Lead acetate cleaves positions G76, A42 and G28, which surround the ribozyme cleavage site. Both Mg2+ and neomycin prevent lead cleavage. Footprinting experiments using base-specific chemical probes revealed enhanced modifications of a set of bases by neomycin, overlapping with the above mentioned lead cleavages. These observations may indicate that neomycin directly displaces divalent metal ions essential for catalysis.
Rogers
J
J
Department of Microbiology and Immunology, University of British Columbia, Vancouver, Canada.
Chang
A H
AH
von Ahsen
U
U
Schroeder
R
R
Davies
J
J
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001483:Base Sequence; D002240:Carbohydrate Sequence; D017931:DNA Primers; D005821:Genetic Techniques; D003698:Hepatitis Delta Virus / Q000201:enzymology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D009355:Neomycin / Q000494:pharmacology*; D016337:RNA, Catalytic / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism
D000900:Anti-Bacterial Agents; D017931:DNA Primers; D016337:RNA, Catalytic; D009355:Neomycin