Functional analysis of reverse transcription by a frameshift pol mutant of murine leukemia virus.
Endogenous reverse transcription by wild-type murine leukemia virus (MuLV) was compared to that catalyzed by clone 23, a pol mutant containing a reverse transcriptase protein which lacks the carboxyl-terminal third of the molecule (J. G. Levin, S. C. Hu, A. Rein, L. I. Messer, and B. I. Gerwin (1984), J. Virol. 51, 470-478). Competition immunoassays revealed that mutant virions contain normal amounts of polymerase protein, indicating that the lack of carboxyl-terminal sequences does not alter normal processing of enzyme precursors. Although the mutant enzyme was previously shown to have the ability to copy and degrade RNA:DNA hybrids, the present study demonstrates that it is defective in functions required to generate full-length copies of viral DNA. Analysis of products of endogenous reverse transcription showed that minus-strand strong-stop DNA is formed and that mutant virions synthesize a series of minus-strand DNA intermediates up to 2.2 kb in length. Comparison of mutant and wild-type MuLV reaction products indicated that the 2.2-kb termination site of the mutant corresponds to a normal pausing region for the wild-type enzyme. Computer analysis of sequences and structure within pausing regions suggested the involvement of C-rich consensus sequences plus multibranch loop structures in the general phenomenon of enzyme-pausing during reverse transcription.
Messer
L I
LI
Currey
K M
KM
O'Neill
B J
BJ
Maizel
J V
JV
Levin
J G
JG
Gerwin
B I
BI
D004279:DNA, Viral / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics*; D005814:Genes, Viral; D009052:Leukemia Virus, Murine / Q000201:enzymology / Q000235:genetics*; D009154:Mutation; D012367:RNA, Viral / Q000235:genetics; D012194:RNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
D004279:DNA, Viral; D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase