Intrinsic DNA synthesis fidelity of xenotropic murine leukemia virus-related virus reverse transcriptase

Abstract

Although recent reports have provided strong evidence to suggest that xenotropic murine leukemia virus‐related virus (XMRV) is unlikely to be the causative agent of prostate cancer and chronic fatigue syndrome, this recombinant retrovirus can nonetheless infect human cells in vitro and induce a chronic infection in macaques. In the present study, we determined the accuracy of DNA synthesis of the reverse transcriptases (RTs) of XMRV and Moloney murine leukemia virus (MoMLV) using a combination of pre‐steady‐state kinetics of nucleotide incorporation and an M13mp2‐based forward mutation assay. The results obtained were compared with those previously reported for the HIV type 1 BH10 strain (HIV‐1 BH10) RT. MoMLV and XMRV RTs were 13.9 and 110 times less efficient [as determined by the catalytic rate constant of the nucleotide incorporation reaction (kpol)/equilibrium constant (Kd)] than the HIV‐1 BH10 RT in incorporating correct nucleotides. Misinsertion and mispair extension kinetic studies demonstrated that MoMLV RT was more accurate than the HIV‐1 BH10 RT. In comparison with the MoMLV RT, the XMRV RT showed decreased mispair extension fidelity and was less faithful when misincorporating C or A opposite A. However, the XMRV RT showed stronger selectivity against G in misinsertion fidelity assays. Forward mutation assays revealed that XMRV and MoMLV RTs had similar accuracy of DNA‐dependent DNA synthesis, but were > 13 times more faithful than the HIV‐1 BH10 enzyme. The mutational spectra of XMRV and MoMLV RTs were similar in having a relatively higher proportion of frameshifts and transversions compared with the HIV‐1 BH10 RT. However, the XMRV polymerase was less prone to introduce large deletions and one‐nucleotide insertions.