Cd. Kelleher et Jj. Champoux, CHARACTERIZATION OF RNA STRAND DISPLACEMENT SYNTHESIS BY MOLONEY MURINE LEUKEMIA-VIRUS REVERSE-TRANSCRIPTASE, The Journal of biological chemistry, 273(16), 1998, pp. 9976-9986
The RNase H activity of reverse transcriptase (RT) is presumably requi
red to cleave the RNA genome following minus strand synthesis to free
the DNA for use as a template during plus strand synthesis. However, s
ince RNA degradation by RNase H appears to generate RNA fragments too
large to spontaneously dissociate from the minus strand, we have inves
tigated the possibility that RNA displacement by RT during plus strand
synthesis contributes to the removal of RNA fragments. By using an RN
ase H- mutant of Moloney murine leukemia virus (M-MuLV) RT, we demonst
rate that the polymerase can displace long regions of RNA in hybrid du
plex with DNA but that this activity is approximately 20-fold slower t
han DNA displacement and 20-fold slower than non-displacement synthesi
s. Furthermore, we find that although certain hybrid sequences seem ne
arly refractory to the initiation of RNA displacement, the same sequen
ces may not significantly impede synthesis when preceded by a single-s
tranded gap. We find that the rate of RNA displacement synthesis by wi
ld-type M-MuLV RT is significantly greater than that of the RNase H- R
T but remains less than the rate of non-displacement synthesis. M-MuLV
nucleocapsid protein increases the rates of RNA and DNA displacement
synthesis approximately 2-fold, and this activity appears to require t
he zinc finger domain.