[Cloning and expression in Escherichia coli of reverse transcriptase coded by the mobile genetic element jockey].
The mobile element jockey is similar in structural organization and coding potential to the LINEs of various organisms. Current models of the mechanism of transposition involve reverse transcription of an RNA intermediate and utilization of element-encoded proteins. As it is demonstrated here, a 2.23 kb DNA fragment from the region of the jockey encoding the putative reverse transcriptase, was stably introduced into the expression system under inducible control of the Escherichia coli lac regulatory elements. We describe the expression of the 92 kDa protein and identify this polypeptide alone as authentic jockey reverse transcriptase based on some of its physical and enzymic properties. The jockey polymerase demonstrates RNA-directed and DNA-directed DNA polymerase activities, but lacks detectable RNase H, has a temperature optimum at 26 degrees C, requires Mg2+ or Mn2+ as a cofactor and is inactivated by sulfhydryl reagent. The enzyme prefers poly(rC) and poly(rA) as template and "activated" DNA is not effective. The results of this work suggest that the RNA-directed DNA polymerase coded by jockey elements may be involved in the transcription of the elements.
Ivanova
V A
VA
Mel'nikov
A A
AA
Siunov
A V
AV
Fodor
I I
II
Il'in
Iu V
IuV
D000595:Amino Acid Sequence; D001483:Base Sequence; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005798:Genes, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D010957:Plasmids; D012194:RNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism
D004251:DNA Transposable Elements; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase; D016914:Ribonuclease H