Hayakawa T, Misumi Y, Kobayashi M, Ohi Y, Fujisawa Y, Kakinuma A, Hatanaka M
Biotechnology Research Laboratories, Takeda Chemical Industries, Ltd., Osaka, Japan.
Biochem Biophys Res Commun. 1991 Dec 31;181(3):1281-7. doi: 10.1016/0006-291x(91)92077-w.
Human T-cell leukemia virus type I (HTLV-I) genome is believed to encode its own protease, although the protease has not yet been detected. To identify the HTLV-I protease, an in-frame gag (3' portion)-prt region was expressed in Escherichia coli. The 14-kDa product was detected using antisera against a synthetic peptide mimicking the fragment of HTLV-I protease, although the molecular weight of the primary translational product was 27,000. A cell extract had a proteolytic activity to cleave a synthetic peptide substrate containing the cleavage site of gag p19/p24 at the correct site in vitro. Replacement of the putative active site Asp-64 with Gly abolished both in vivo processing activity and in vitro proteolytic activity. These results suggest that the 14-kDa product is the mature enzymatically active HTLV-I protease generated through posttranslational autoprocessing in E. coli.
人们认为人类嗜T细胞病毒I型（HTLV-I）基因组编码自身的蛋白酶，尽管尚未检测到该蛋白酶。为了鉴定HTLV-I蛋白酶，在大肠杆菌中表达了一个读码框内的gag（3'部分）-prt区域。使用针对模拟HTLV-I蛋白酶片段的合成肽的抗血清检测到了14 kDa的产物，尽管初级翻译产物的分子量为27,000。细胞提取物在体外具有蛋白水解活性，可在正确位点切割包含gag p19/p24切割位点的合成肽底物。将推定的活性位点天冬氨酸-64替换为甘氨酸消除了体内加工活性和体外蛋白水解活性。这些结果表明，14 kDa的产物是通过在大肠杆菌中进行翻译后自加工产生的成熟的具有酶活性的HTLV-I蛋白酶。
