Ladner S K, Miller T J, Otto M J, King R W
Avid Therapeutics, Philadelphia, PA 19104, USA.
Antivir Chem Chemother. 1998 Jan;9(1):65-72.
A variant of hepatitis B virus (HBV) containing a Met-to-Val substitution (M539V) in the YMDD motif of the polymerase nucleoside-binding domain exhibited resistance to the cytosine analogue lamivudine (3TC). To determine if the mutation responsible for the M539V polymerase variant affected the sensitivity of the virus to other nucleoside analogues, we constructed a tetracycline-responsive cell line, HepAD79. This cell line is stably transfected with a cDNA copy of the pregenomic RNA of an HBV genome containing an A-to-G mutation in the first position of the polymerase gene codon 539. This mutation results in a Met-to-Val substitution at amino acid 539 of the polymerase. When grown under the proper conditions, HepAD79 cells produced HBV RNA, contained HBV DNA associated with immature core particles and released core-associated HBV DNA into the medium. The M539V polymerase variant produced in this cell line was approximately 26-fold less sensitive to the antiviral effects of 3TC than wild-type virus. In addition, this variant demonstrated decreased sensitivity to the cytosine analogues FTC and ddC, as well as the thymidine analogue AZT.
在聚合酶核苷结合域的YMDD基序中含有甲硫氨酸至缬氨酸替代（M539V）的乙型肝炎病毒（HBV）变体对胞嘧啶类似物拉米夫定（3TC）表现出耐药性。为了确定导致M539V聚合酶变体的突变是否影响病毒对其他核苷类似物的敏感性，我们构建了一种四环素反应性细胞系HepAD79。该细胞系用含有聚合酶基因密码子539第一位A到G突变的HBV基因组前基因组RNA的cDNA拷贝进行稳定转染。该突变导致聚合酶氨基酸539位的甲硫氨酸至缬氨酸替代。在适当条件下培养时，HepAD79细胞产生HBV RNA，含有与未成熟核心颗粒相关的HBV DNA，并将与核心相关的HBV DNA释放到培养基中。在该细胞系中产生的M539V聚合酶变体对3TC抗病毒作用的敏感性比野生型病毒低约26倍。此外，该变体对胞嘧啶类似物FTC和ddC以及胸苷类似物AZT的敏感性降低。
