JOURNALOFVIROLOGY,June1999,p.PDFVIP

JOURNAL OF VIROLOGY, June 1999, p. 4794–4805 Vol. 73, No. 6 0022-538X/99/$04.00?0 Copyright ? 1999, American Society for Microbiology. All Rights Reserved. Molecular Requirements for Human Immunode?ciency Virus Type 1 Plus-Strand Transfer: Analysis in Reconstituted and Endogenous Reverse Transcription Systems 1 1 2 2 1 TIYUN WU, JIANHUI GUO, JULIAN BESS, LOUIS E. HENDERSON, AND JUDITH G. LEVIN * Laboratory of Molecular Genetics, National Institute of Child Health and Human Development, Bethesda, Maryland 20892,1 and AIDS Vaccine Program, SAIC-Frederick, NCI-Frederick Cancer Research and Development Center, Frederick, Maryland 217022 Received 29 December 1998/Accepted 9 March 1999 We have developed a reconstituted system which models the events associated with human immunode?ciency virus type 1 (HIV-1) plus-strand transfer. These events include synthesis of plus-strand strong-stop DNA [(?) Downloaded from Lys SSDNA] from a minus-strand DNA donor template covalently attached to human tRNA3 , tRNA primer (removal, and annealing of (?) SSDNA to the minus-strand DNA acceptor template. Termination of (? ? Lys SSDNA synthesis at the methyl A (nucleotide 58) near the 3 end of tRNA3 reconstitutes the 18-nucleotide primer binding site (PBS). Analysis of (?) SSDNA synthesis in vitro and in HIV-1 endogenous reactions indicated another major termination site: the pseudouridine at nucleotide 55. In cer