Expression and characterization of a novel reverse transcriptase of the LTR retrotransposon Tf1.
The LTR retrotransposon of Schizosacharomyces pombe, Tf1, has several distinctive properties that can be related to the unique properties of its reverse transcriptase (RT). Consequently, we expressed, purified and studied the recombinant Tf1 RT. This monomeric protein possesses all activities typical to RTs: DNA and RNA-dependent DNA polymerase as well as an inherent ribonuclease H. The DNA polymerase activity shows preference to Mn(+)(2) or Mg(+)(2), depending on the substrate used, whereas the ribonuclease H strongly prefers Mn(+)(2). The most outstanding feature of Tf1 RT is its capacity to add non-templated nucleotides to the 3'-ends of the nascent DNA. This is mainly apparent in the presence of Mn(+)(2), as is the noticeable low fidelity of DNA synthesis. In all, Tf1 RT has a marked infidelity in synthesizing DNA at template ends, a phenomenon that can explain, as discussed herein, some of the features of Tf1 replication in the host cells.
Kirshenboim
Noa
N
Department of Cell and Developmental Biology, Sackler School of Medicine, Tel-Aviv University, Tel-Aviv, 69978, Israel.
Hayouka
Zvi
Z
Friedler
Assaf
A
Hizi
Amnon
A
D003001:Cloning, Molecular; D004247:DNA / Q000096:biosynthesis; D020536:Enzyme Activators / Q000494:pharmacology; D015870:Gene Expression; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D009711:Nucleotides / Q000378:metabolism; D021122:Protein Subunits; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D018626:Retroelements / Q000235:genetics*; D016914:Ribonuclease H / Q000378:metabolism; D012568:Schizosaccharomyces / Q000201:enzymology* / Q000235:genetics
D020536:Enzyme Activators; D009711:Nucleotides; D021122:Protein Subunits; D018626:Retroelements; D008345:Manganese; D004247:DNA; D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H; D008274:Magnesium