The non-LTR (long terminal repeat) retrotransposon L1Tc from Trypanosoma cruzi codes for a protein with RNase H activity

Mónica Olivares, José Luis García-Pérez, M Carmen Thomas, Sara R Heras, Manuel C López

Research output: Contribution to journalArticlepeer-review

Abstract

The deduced amino acid sequence of the region downstream of the reverse transcriptase (RT) motif of the Trypanosoma cruzi L1Tc non-LTR retrotransposon shows a significant homology with the sequence coding for proteins with RNase H activity from different organisms and retroelements. The 25-kDa His(6)-tagged recombinant protein bearing only the L1Tc RNase H domain, named RHL1Tc, exhibits RNase H activity as measured on the [(3)H]poly(rA)/poly(dT) hybrid used as substrate as well as on specific homologous and heterologous [(32)P]RNA/DNA hybrids. The mutation of the conserved aspartic acid at position 39 of the enzyme catalytic site, but not of the serine at position 56 (non-conservative amino acid), abolishes protein RNase H activity. The RNase H activity of the RHL1Tc protein is Mg(2+)-dependent, and it is also active in the presence of the Mn(2+) ion. The optimal condition of RNase H activity is found at pH 8 and 37 degrees C, although it also has significant enzymatic activity at 19 degrees C and pH 6. However, it cannot be excluded that the RNase H activity level and its optimal conditions may be different from that of a protein containing both RT and RNase H domains.

Original languageEnglish
Pages (from-to)28025-30
Number of pages6
JournalJournal of Biological Chemistry
Volume277
Issue number31
DOIs
Publication statusPublished - 2 Aug 2002

Keywords

  • Amino Acid Sequence
  • Animals
  • Consensus Sequence
  • DNA, Protozoan
  • Kinetics
  • Long Interspersed Nucleotide Elements
  • Molecular Sequence Data
  • Nucleic Acid Hybridization
  • RNA, Protozoan
  • RNA-Directed DNA Polymerase
  • Retroelements
  • Ribonuclease H
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Thermodynamics
  • Trypanosoma cruzi

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