Specificity of priming reaction of HIV-1 reverse transcriptase, 2'-OH or 3'-OH.
It has not been unambiguously demonstrated whether the priming reaction of human immunodeficiency virus, type 1 (HIV-1) cDNA synthesis initiates with either the 2'-OH or 3'-OH group of the 3'-terminal adenosine residue of tRNA(Lys-3). In this report, we synthesized tRNA(Lys-3) of which the 3'-terminal adenosine residue lacks either a 2'-OH or 3'-OH. These tRNA molecules were used for the HIV-1 cDNA-priming reaction in a cell-free system consisting of a 141-base RNA template and purified HIV-1 reverse transcriptase. It was found that under the conditions used, the tRNA containing the 2'-deoxyadenosine was able to initiate the cDNA synthesis, while the tRNA with the 3'-deoxyadenosine was not. The results show that retroviral reverse transcriptase specifically primes cDNA synthesis from the 3'-OH group. This is in contrast to bacterial reverse transcriptase, which initiates cDNA synthesis from the 2'-OH group of an internal guanosine residue of a template RNA.
Shimada
M
M
Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854.
Hosaka
H
H
Takaku
H
H
Smith
J S
JS
Roth
M J
MJ
Inouye
S
S
Inouye
M
M
D001483:Base Sequence; D002474:Cell-Free System; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000096:biosynthesis; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology; D008969:Molecular Sequence Data; D012357:RNA, Transfer, Lys / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D013379:Substrate Specificity
D017931:DNA Primers; D018076:DNA, Complementary; D012357:RNA, Transfer, Lys; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase