The p51 subunit of human immunodeficiency virus type 1 reverse transcriptase is essential in loading the p66 subunit on the template primer.
Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is a dimeric enzyme consisting of p66 and p51 subunits. The functional role of the p51 subunit remains elusive since all the catalytic functions appear to be executed through the p66 subunit. We report here that the p51 subunit, in addition to providing structural support to the p66 subunit, may be involved in facilitating the loading of the p66 subunit on to the template-primer (TP). This possibility is supported by following observations: (i) Upon binding to the TP, the p51 subunit can be dissociated by acetonitrile treatment and the template-primer-bound p66 monomer alone is capable of catalyzing DNA synthesis. (ii) Photo-cross-linking of template-primer to HIV-1 RT is abolished by dissociation of the p51 subunit prior to the TP binding but remains unaffected after the TP binding step. (iii) The p66-TP covalent complex selectively generated by UV irradiation and separated by gel electrophoresis can incorporate a single nucleotide in situ upon its renaturation in the gel. (iv) Treatment of HIV-1 RT with (tert-butyldimethylsilyl)spiroaminooxathioledioside (TSAO), an inhibitor that specifically binds to the beta7 beta8 loop of p51, destabilizes the heterodimeric enzyme, resulting in the subsequent loss of DNA binding.
Harris
D
D
Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry at New Jersey Medical School, Newark 07103, USA.
Lee
R
R
Misra
H S
HS
Pandey
P K
PK
Pandey
V N
VN
D000097:Acetonitriles; D001483:Base Sequence; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D004256:DNA Polymerase I / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D019281:Dimerization; D054303:HIV Reverse Transcriptase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000235:genetics; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D013698:Templates, Genetic
D000097:Acetonitriles; D017931:DNA Primers; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; D004256:DNA Polymerase I; D004259:DNA-Directed DNA Polymerase; C032159:acetonitrile