Modulation of HIV-1 reverse transcriptase function in "selectively deleted" p66/p51 heterodimers

Area of Science:

• Molecular Biology
• Virology
• Biochemistry

Background:

• Human immunodeficiency virus type-1 (HIV-1) reverse transcriptase (RT) is a key enzyme for viral replication.
• The heterodimeric RT (p66/p51) possesses polymerase and ribonuclease H (RNase H) activities.
• The precise roles of the p51 subunit in RT function, particularly primer interaction, require further elucidation.

Purpose of the Study:

• To investigate the contribution of the p51 subunit of HIV-1 RT to heterodimer activities.
• To assess the impact of C-terminal deletions in the p51 subunit on RT function and primer binding.
• To determine the specific role of p51 in the interaction with the HIV-1 replication primer, tRNA(Lys,3).

Main Methods:

• Construction and reconstitution of selectively deleted heterodimers of HIV-1 RT.
• Assay of polymerase and RNase H enzymatic activities.
• Analysis of the interaction between reconstituted heterodimers and the tRNA(Lys,3) primer.

Main Results:

• C-terminal truncations of the p51 subunit modulated the reconstitution efficiency and enzymatic functions of the heterodimer.
• A p51 subunit lacking 13 C-terminal residues resulted in severely impaired tRNA(Lys,3) binding.
• Despite impaired tRNA binding, enzymatic activities (polymerase and RNase H) were retained in the p51-deleted mutant.

Conclusions:

• The extent of C-terminal deletion in the p51 subunit influences HIV-1 RT heterodimer properties.
• The p51 subunit is critically involved in the binding of the tRNA(Lys,3) primer.
• These findings support a significant role for the p51 subunit in the initiation of HIV-1 reverse transcription.