Insight into the Mechanism of the Stabilization of Moloney Murine Leukaemia Virus Reverse Transcriptase by Eliminating RNase H Activity
Insight into the Mechanism of the Stabilization of Moloney Murine Leukaemia Virus Reverse Transcriptase by Eliminating RNase H Activity
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DOI：
10.1271/bbb.90777
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发表时间：
2010-02-01
影响因子：
1.6
通讯作者：
Inouye, Kuniyo
中科院分区：
文献类型：
--
作者：
Mizuno, Masaki;Yasukawa, Kiyoshi;Inouye, Kuniyo
我们探索了通过消除 RNase H 活性来稳定莫洛尼鼠白血病病毒逆转录酶 (MMLV RT) 的机制。在没有模板引物 (T/P) poly(rA)-p(dT)(15) 的情况下，RNase H 活性缺陷变体 D524A 在 10 分钟的孵育过程中，初始逆转录活性降低 50% 的温度比野生型酶 (WT) 高 3.7 摄氏度。在逆转录反应中，WT和D524A的T/P的K-m值几乎相同。这些结果表明，RNase H 活性的消除增强了 MMLV RT 的内在热稳定性，而不是其对 T/P 的亲和力。
We explored the mechanism of the stabilization of Moloney murine leukaemia virus reverse transcriptase (MMLV RT) by eliminating RNase H activity. Without the template-primer (T/P) poly(rA)-p(dT)(15), the temperature reducing initial reverse-transcription activity by 50% over a 10-min incubation of the RNase H activity-deficient variant D524A was higher by 3.7 degrees C than that of the wild-type enzyme (WT). In the reverse transcription reaction, the K-m values for T/P of WT and D524A were almost the same. These results suggest that elimination of RNase H activity enhanced the intrinsic thermal stability of MMLV RT rather than its affinity toward T/P.