THE DIFFERENTIAL PROCESSING OF HOMODIMERS OF REVERSE TRANSCRIPTASES FROM HUMAN IMMUNODEFICIENCY VIRUSES TYPE-1 AND TYPE-2 IS A CONSEQUENCE OF THE DISTINCT SPECIFICITIES OF THE VIRAL PROTEASES

Active, recombinant p68 reverse transcriptase (RT) from human immunode ficiency virus type 2 (HIV-2), with an NH2-terminal extension containi ng a hexahistidine sequence was isolated from extracts of Escherichia coli by immobilized metal affinity chromatography. Treatment of the pu rified p68/p68 homodimer of HIV-2 RT with recombinant HIV-2 protease g enerates stable, active heterodimer (p68/p58) that is resistant to fur ther hydrolysis. Analysis of this p68/p58 HIV-2 RT heterodimer reveale d that while one subunit is intact p68, the p58 subunit is COOP-termin ally truncated by cleavage, not at Phe(440) as is seen in processing o f the p66/p66 HIV-1 RT homodimer by HIV-1 protease, but at Met(484) Th e expected COOP-terminal p10 fragment resulting from hydrolysis of p68 at Met(484) is not released intact, but undergoes further cleavage at Asn(494), Met(503), and Tyr(532). Processing of p68/p68 HlV-2 RT with the HIV-1 protease led to cleavage of the phe(440)-Tyr(441) bond, exa ctly as is seen with p66/p66 HTV-1 RT, to give the analogous p53 subun it. Studies of a peptide substrate modeled after residues 437-444 in H IV-2 RT showed that while the HIV-1 protease was able to cleave the Ph e(440) bond, this bond was resistant to cleavage by the HIV-2 enzyme. Our findings provide a rationale for the previous observation that the RT heterodimer isolated from HIV-P lysates is larger than that from H IV-1. We conclude that the p68/p58 HIV-2 RT heterodimer, containing th e Met(484) truncated p58 subunit, is a biologically relevant form of t he enzyme in vivo.