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J. Biol. Chem., Vol. 269, Issue 6, 3925-3927, 02, 1994

Specificity of priming reaction of HIV-1 reverse transcriptase, 2'-OH or 3'-OH

M Shimada, H Hosaka, H Takaku, JS Smith, MJ Roth, S Inouye and M Inouye
Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854.

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.
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				T. Shimamoto, M. Inouye, and S. Inouye The Formation of the 2`,5`-Phosphodiester Linkage in the cDNA Priming Reaction by Bacterial Reverse Transcriptase in a Cell-free System J. Biol. Chem., January 13, 1995; 270(2): 581 - 588. [Abstract] [Full Text] [PDF]