Document 0162
 DOCN  M9650162
 TI    Helix structure and ends of RNA/DNA hybrids direct the cleavage
       specificity of HIV-1 reverse transcriptase RNase H.
 DT    9605
 AU    Palaniappan C; Fuentes GM; Rodriguez-Rodriguez L; Fay PJ; Bambara RA;
       Department of Biochemistry, University of Rochester, New York; 14642,
       USA.
 SO    J Biol Chem. 1996 Jan 26;271(4):2063-70. Unique Identifier : AIDSLINE
       MED/96147182
 AB    RNA/DNA hybrids in human immunodeficiency virus (HIV) replication are
       cleaved by HIV-1 reverse transcriptase (RT) H in locations determined by
       hybrid structure. Minus strand DNA synthesis is accompanied by cleavage
       of template viral RNA directed by RT positioned at the growing 3' DNA
       end. Some RNA remains as oligomers annealed to the new DNA strand and is
       cut by RTs positioned at the 5' RNA ends. We constructed substrates to
       the test the hypothesis that internal helix structure, rather than
       strand end structure, drives the RT to position at 3' DNA and 5' RNA
       ends. On substrates with an RNA primer recessed on a DNA template, the
       5' end of the RNA had a dominant role in the determination of RNase H
       cleavage positions. If the 5' end region of the RNA could not anneal,
       cleavage would not occur. Nevertheless, we obtained evidence that helix
       structure promotes the binding of RT to the end of the helical region
       closest to the 5' RNA/3' DNA end. When a DNA primer recessed on an RNA
       template had a 3' unannealed region, cleavage occurred, with RT
       positioned solely by helical structure at the 5' RNA/3' DNA end of the
       annealed region of the hybrid. Using substrates having RNA primers
       annealed to circular DNA templates, we showed that cleavage can be
       independent of the presence of a DNA 3'end and is directed by the 5' RNA
       end. Overall, the results suggest that the RT initially binds an
       internal region of the hybrid and then is driven in the direction to
       encounter a 3' DNA or 5' RNA end, where it is positioned for catalysts
       by the strand end. The requirement for two modes of RNA cleavage in
       viral replication and the unexpected requirement for the 5' RNA end
       structure are discussed.
 DE    Base Sequence  DNA Primers/CHEMISTRY  DNA, Single-Stranded/METABOLISM
       DNA, Viral/METABOLISM  Kinetics  Molecular Sequence Data  Nucleic Acid
       Hybridization  Recombinant Proteins  Ribonuclease H, Calf
       Thymus/CHEMISTRY/*METABOLISM  RNA-Directed DNA
       Polymerase/CHEMISTRY/*METABOLISM  RNA, Viral/METABOLISM
       Structure-Activity Relationship  Substrate Specificity  Support, U.S.
       Gov't, P.H.S.  Templates  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).