Insight into the mechanism of the stabilization of moloney murine leukaemia virus reverse transcriptase by eliminating RNase H activity

Biosci Biotechnol Biochem. 2010;74(2):440-2. doi: 10.1271/bbb.90777. Epub 2010 Feb 7.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Enzyme Stability
  • Kinetics
  • Moloney murine leukemia virus / enzymology*
  • Mutation*
  • RNA-Directed DNA Polymerase / genetics
  • RNA-Directed DNA Polymerase / metabolism*
  • Reverse Transcription
  • Ribonuclease H / genetics*
  • Ribonuclease H / metabolism*
  • Temperature

Substances

  • RNA-Directed DNA Polymerase
  • Ribonuclease H