Abstract
The interaction of several forms (p51, p66, and p66/p51) of recombinant human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) with a synthetic derivative of its cognate replication primer, tRNA(Lys-3), has been determined by gel-mobility shift analysis. While p66/p51 RT is proficient in tRNA binding, preparations of p66 and p51 display only weak binding at elevated protein:tRNA ratios, despite the former containing both RNA-dependent DNA polymerase and ribonuclease H (RNase H) activity. Gel permeation analysis of purified p66 RT indicate this to be predominantly monomeric, suggesting that dimerization may be a prerequisite for efficient tRNA binding. Prolonged incubation of a mixture of the 66- and 51-kDa polypeptides results in heterodimer reconstitution, restoration of tRNA binding, and recovery of appreciable levels of RNA-dependent DNA polymerase activity. Under the same conditions, both the tRNA binding and RNA-dependent DNA polymerase activities of the 66- and 51-kDa polypeptides are unaffected, suggesting that they remain in the monomeric conformation.
Highlights
From the $Department of Biochemistry and the §Divisioof nInfectious Diseases, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106
Alwould predict 0.2 pmol of dimer, which might account for the weak tRNA binding observed at thipsrotein:tRNA ratio
Despite displaying RNA-dependent DNA polymactivity, there is insufficient evidence to determine whether erase and RNase H activity, as well as a capacity for reconthese can be attributedto monomeric or dimeric protein. stitution with the 51-kDa HIV-1 polypeptide, the chimeric
Summary
From the $Department of Biochemistry and the §Divisioof nInfectious Diseases, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106. Prolonged incubation of a mixture of the66- and 51- We (11, 12) and others (13) have demonstrated that the kDa polypeptides results in heterodimerreconstitu- p66/p51 form of HIV-1 RT forms a stable and specific comtion, restorationof tRNA binding,andrecoveryof plex with either the natural oirn vitro-synthesized version of appreciablelevels of RNA-dependent DNA polymeraseits cognate replication primer, tRNALy"-3I,n both cases, the activity.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
