title: The protease and reverse transcriptase of the tobacco LTR retrotransposon Tnt1 are enzymatically active when expressed in Escherichia coli.
abstract: The open reading frame (ORF) of the tobacco retrotransposon Tnt1-94 was over-expressed in Escherichia coli to assay its protease and reverse transcriptase (RT) enzymatic activities. In E. coli, Tnt1-94 polyprotein is cleaved off by the element-encoded protease to release a Gag protein with an apparent molecular mass of 37 kDa that forms high-density aggregates. The catalytic site of Tnt1-94 protease (D-T-A) as determined by deletion analysis differs from that of retroviruses and of well-characterized retrotransposons (D-T/S-G). The cleaved or uncleaved ORF of Tnt1-94 displays an exogenous RT activity. Over-expression of plant retrotransposons ORFs in E. coli provides a very useful strategy to assay the enzymatic activities of their proteins and to determine their catalytic sites.
authors.affiliation: Institut National de la Recherche Agronomique, Laboratoire de Biologie Cellulaire, Versailles, France.
authors.forename: F
authors.initials: F
authors.lastname: Feuerbach
authors.forename: H
authors.initials: H
authors.lastname: Lucas
mesh_terms: D015153:Blotting, Western; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D010947:Plants, Toxic; D012194:RNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D018626:Retroelements / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; D018626:Retroelements; D012194:RNA-Directed DNA Polymerase; D010450:Endopeptidases