Operario Darwin J, Reynolds Holly M, Kim Baek
Department of Microbiology and Immunology, School of Medicine and Dentistry, University of Rochester, 601 Elmwood Avenue, Box 672, Rochester, NY 14642, USA.
Virology. 2005 Apr 25;335(1):106-21. doi: 10.1016/j.virol.2005.02.010.
In this study, we present enzymatic differences found between recombinant RTs derived from feline leukemia virus and feline immunodeficiency virus. Firstly, FIV RT showed low steady state K(m) values for dNTPs compared to FeLV RT. Consistent with this, FIV RT synthesized DNA more efficiently than FeLV RT at low dNTP concentrations. We observed similar concentration-dependent activity differences between other lentiviral (HIV-1 and SIV) and non-lentiviral (MuLV and AMV) RTs. Second, FeLV RT showed less efficient misincorporation with biased dNTP pools and mismatch primer extension capabilities, compared to FIV RT. In M13mp2 lacZalpha forward mutation assays, FeLV RT displayed approximately 11-fold higher fidelity than FIV RT. Finally, FeLV RT was less sensitive to 3TCTP and ddATP than FIV RT. This study represents the comprehensive enzymatic characterization of RTs from a lentivirus and a non-lentivirus retrovirus from the same host species. The data presented here support enzymatic divergences seen among retroviral RTs.
在本研究中,我们展示了源自猫白血病病毒和猫免疫缺陷病毒的重组逆转录酶(RT)之间发现的酶学差异。首先,与猫白血病病毒RT相比,猫免疫缺陷病毒RT对脱氧核苷三磷酸(dNTP)的稳态米氏常数(K(m))值较低。与此一致的是,在低dNTP浓度下,猫免疫缺陷病毒RT比猫白血病病毒RT更有效地合成DNA。我们在其他慢病毒(HIV-1和SIV)和非慢病毒(鼠白血病病毒和禽成髓细胞瘤病毒)RT之间观察到了类似的浓度依赖性活性差异。其次,与猫免疫缺陷病毒RT相比,猫白血病病毒RT在dNTP库有偏差时错配掺入效率较低,且错配引物延伸能力较弱。在M13mp2 lacZα正向突变试验中,猫白血病病毒RT的保真度比猫免疫缺陷病毒RT高约11倍。最后,猫白血病病毒RT对3'-硫代胞苷三磷酸(3TCTP)和双脱氧腺苷三磷酸(ddATP)的敏感性低于猫免疫缺陷病毒RT。本研究代表了来自同一宿主物种的慢病毒和非慢病毒逆转录病毒RT的全面酶学特征。此处呈现的数据支持了逆转录病毒RT之间存在的酶学差异。