Alternative modes of polymerization distinguish the subunits of equine infectious anemia virus reverse transcriptase.
A comparative study of recombinant 51- and 66-kDa subunits comprising equine infectious anemia virus reverse transcriptase (EIAV RT) is reported. Both polypeptides sedimented as stable homodimers (molecular mass, 102 and 132 kDa, respectively) when analyzed by rate sedimentation through glycerol gradients. Consistent with their dimer composition, each preparation displayed considerable levels of both RNA- and DNA-dependent DNA polymerase activity on different homopolymeric template/primer combinations. However, a detailed analysis of the polymerization products indicated qualitative differences. Whereas p66 EIAV RT proceeded essentially unimpaired along both RNA and DNA templates, p51-catalyzed DNA synthesis was interrupted close to or in the immediate vicinity of the primer. A series of "programmed" 2-step polymerization reactions suggests that p51 EIAV RT enters an abortive mode of polymerization. Duplication of this observation with p51 human immunodeficiency virus-1 RT, together with recent observations from murine RT, suggests that lack of a ribonuclease H domain and loss of contact with the nascent product from the polymerase active center have profound consequences on the mode of polymerization.
Wöhrl
B M
BM
Division of Infectious Diseases, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
Howard
K J
KJ
Jacques
P S
PS
Le Grice
S F
SF
D001483:Base Sequence; D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D054303:HIV Reverse Transcriptase; D006801:Humans; D004860:Infectious Anemia Virus, Equine / Q000201:enzymology*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013698:Templates, Genetic
D017931:DNA Primers; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase