Document 0333
 DOCN  M9650333
 TI    Ribonuclease H activity during initiation of reverse transcription using
       tRNA(lys)/RNA primer/template of human immunodeficiency virus.
 DT    9605
 AU    Artzi HB; Shemesh J; Zeelon E; Amit B; Kleiman L; Gorecki M; Panet A;
       Bio-Technology General Israel, Ltd., Kiryat Weizmann, Rehovot,; Israel.
 SO    Arch Biochem Biophys. 1996 Jan 15;325(2):209-16. Unique Identifier :
       AIDSLINE MED/96139301
 AB    The specificity of the initial cleavage by the RNaseH activity of HIV-1
       reverse transcriptase (RT) during minus strong-stop DNA synthesis was
       studied using the authentic primer/template tRNA(lys)/HIV RNA. We
       observed that concomitant with the initiation of DNA synthesis, RNaseH
       activity of HIV RT introduced the first endonucleolytic cuts within the
       U5 region of the HIV RNA template, mainly 1 and 3 bases away from the
       primer binding site. To analyze whether the cleavage sites were
       determined by sequence specificity, the authentic U5 region at one of
       the cleavage sites was mutated. The change of sequence did not alter the
       initial cleavage pattern of RNaseH. In order to determine the size of
       the RNA/DNA hybrid that is required for RNaseH activation during reverse
       transcription initiation, DNA synthesis was limited by
       dideoxynucleotides. DNA extension of the tRNA(lys) primer by 17
       deoxyribonucleotides but not by 6 deoxyribonucleotides was sufficient to
       activate the RNaseH site of HIV RT. Taken together, our results indicate
       that during initiation of minus strongstop DNA synthesis by HIV RT, the
       first RNaseH-mediated endonucleolytic cut of the genomic RNA is dictated
       mainly by the length of the nascent DNA and not by sequence preference.
 DE    Base Sequence  Binding Sites  DNA, Viral/BIOSYNTHESIS/GENETICS  Enzyme
       Activation  Human  HIV-1/GENETICS/*METABOLISM  In Vitro  Molecular
       Sequence Data  Ribonuclease H, Calf Thymus/*METABOLISM  RNA/*GENETICS
       RNA-Directed DNA Polymerase/*METABOLISM  RNA, Transfer,
       Lys/GENETICS/*METABOLISM  RNA, Viral/CHEMISTRY/GENETICS/*METABOLISM
       Support, U.S. Gov't, P.H.S.  Transcription, Genetic  JOURNAL ARTICLE

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