Document 0780
 DOCN  M9590780
 TI    Effects on DNA synthesis and translocation caused by mutations in the
       RNase H domain of Moloney murine leukemia virus reverse transcriptase.
 DT    9509
 AU    Blain SW; Goff SP; Howard Hughes Medical Institute, Department of
       Biochemistry and; Molecular Biophysics, College of Physicians and
       Surgeons,; Columbia University, New York, New York 10032, USA.
 SO    J Virol. 1995 Jul;69(7):4440-52. Unique Identifier : AIDSLINE
       MED/95287501
 AB    To determine the various roles of RNase H in reverse transcription, we
       generated a panel of mutations in the RNase H domain of Moloney murine
       leukemia virus reverse transcriptase based on sequence alignments and
       the crystal structures of Escherichia coli and human immunodeficiency
       virus type 1 RNases H (S. W. Blain and S. P. Goff, J. Biol. Chem.
       268:23585-23592, 1993). These mutations were introduced into a
       full-length provirus, and the resulting genomes were tested for
       infectivity by transient transfection assays or after generation of
       stable producer lines. Several of the mutant viruses replicated
       normally, some showed significant delays in infectivity, and others were
       noninfectious. Virions were collected, and the products of the
       endogenous reverse transcription reaction were examined to determine
       which steps might be affected by these mutations. Some mutants left
       their minus-strand strong-stop DNA in RNA-DNA hybrid form, in a manner
       similar to that of RNase H null mutants. Some mutants showed increased
       polymerase pausing. Others were impaired in first-strand translocation,
       independently of their wild-type ability to degrade genomic RNA,
       suggesting a new role for RNase H in strand transfer. DNA products
       synthesized in vivo by the wild-type and mutant viruses were also
       examined. Whereas wild-type virus did not accumulate detectable levels
       of minus-strand strong-stop DNA, several mutants were blocked in
       translocation and did accumulate this intermediate. These results
       suggest that in vivo wild-type virus normally translocates minus-strand
       strong-stop DNA efficiently.
 DE    Animal  Base Sequence  Biological Transport  DNA, Viral/*METABOLISM
       Mice  Molecular Sequence Data  Moloney Leukemia Virus/*ENZYMOLOGY
       Mutation  Reverse Transcriptase/*PHYSIOLOGY  Ribonuclease H, Calf
       Thymus/*PHYSIOLOGY  RNA, Viral/METABOLISM  Structure-Activity
       Relationship  Support, Non-U.S. Gov't  Support, U.S. Gov't, P.H.S.
       Viral Proteins/ANALYSIS  Virion/CHEMISTRY  Virus Replication  3T3 Cells
       JOURNAL ARTICLE

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