TRUNCATING ALPHA-HELIX E' OF P66 HUMAN-IMMUNODEFICIENCY-VIRUS REVERSE-TRANSCRIPTASE MODULATES RNASE-H FUNCTION AND IMPAIRS DNA STRAND TRANSFER

The properties of recombinant p66/p51 human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) containing C-terminal truncations in its p66 polypeptide were evaluated, Deletion end points partly or completely removed alpha-helix E' of the RNase H domain (p66 Delta 8/p51 and p66 Delta 16/p51, respectively), while mutant p66 Delta 23/p51 lacked alpha E' and the beta 5'-alpha E' connecting loop. Although dimerization and DNA polymerase properties of all mutants were not significantly different from those of the parental enzyme, p66 Delta 16/p51 and p66 Delta 23/p51 RT lacked ribonuclease H (RNase H) activity, In contrast, RT mutant p66 Delta 8/p51 retained endonuclease activity but lacked the directional processing feature of the parental enzyme, Despite retaining full endoribonuclease function, p66 Delta 8/p51 RT barely supported transfer of nascent (-)-strand DNA between RNA templates representing the 5' and 3' ends of retroviral genome, shedding light on the requirement for the endonuclease and directional processing functions of the RNase H domain during replication.