Influence of template primary structure on 3'-azido-3'-deoxythymidine triphosphate incorporation into DNA.
In the present study, templates containing a specific segment of the G gamma-globin gene were constructed and incorporation of 3'-azido-3'-deoxythymidine 5'-triphosphate (AZT-TP) or 2',3'-dideoxythymidine 5'-triphosphate (ddTTP) into these templates was compared to that observed in M13 bacteriophage DNA. Investigations on the intrinsic fidelity of T7 DNA polymerase in reactions with AZT-TP and without deoxythymidine 5'-triphosphate, resulted in DNA synthesis beyond the first T site, suggesting that other normal deoxynucleotides misincorporated at these T sites. Modified T7 DNA polymerase incorporated AZT-TP into T sites of elongating DNA strands. Chain termination at noncomplementary sites was also observed with AZT-TP when a genomic DNA template was used and interestingly, this phenomenon was not detected in the presence of a M13 DNA template. These DNA template-dependent effects were not detected with either ddTTP, 2',3'-dideoxycytidine 5'-triphosphate, or 2',3'-didehydro-2',3'-dideoxythymidine 5'-triphosphate (D4T-TP). A variation in the extent of chain termination at T sites was observed with D4T-TP suggesting that each 2',3'-dideoxynucleoside may exhibit unique chain termination patterns along with the template sequence.
Bridges
E G
EG
Department of Pharmacology, University of Alabama, Birmingham 35294.
LeBoeuf
R B
RB
Weidner
D A
DA
Sommadossi
J P
JP
D000998:Antiviral Agents / Q000378:metabolism*; D017104:Bacteriophage M13; D001483:Base Sequence; D004247:DNA / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D054306:Dideoxynucleotides; D005914:Globins / Q000235:genetics; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D013942:Thymine Nucleotides / Q000378:metabolism*; D015215:Zidovudine / Q000031:analogs & derivatives* / Q000378:metabolism
D000998:Antiviral Agents; D004279:DNA, Viral; D054306:Dideoxynucleotides; D013942:Thymine Nucleotides; D015215:Zidovudine; C052873:zidovudine triphosphate; D005914:Globins; D004247:DNA; C034970:bacteriophage T7 induced DNA polymerase; D004259:DNA-Directed DNA Polymerase