Semizarov D G, Arzumanov A A, Dyatkina N B, Meyer A, Vichier-Guerre S, Gosselin G, Rayner B, Imbach J L, Krayevsky A A
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 32 Vavilov Street, Moscow, 117984 Russia.
J Biol Chem. 1997 Apr 4;272(14):9556-60. doi: 10.1074/jbc.272.14.9556.
All four possible stereoisomers of dNTP with regard to deoxyribofuranose C-1' and C-4' carbon atoms were studied as substrates for several template-dependent DNA polymerases and template-independent terminal deoxynucleotidyl transferase. It was shown that DNA polymerases alpha, beta, and epsilon from human placenta and reverse transcriptases of human immunodeficiency virus and avian myeloblastosis virus incorporate into the DNA chain only natural beta-D-dNTPs, whereas calf thymus terminal deoxynucleotidyl transferase incorporates two nucleotide residues of alpha-D-dNTP and extends the resulting oligonucleotide in the presence of beta-D-dNTPs. The latter enzyme also extended alpha-anomeric D-oligodeoxynucleotide primers in the presence of beta-D-dNTPs. None of the studied enzymes utilized L-dNTPs. These data indicate that template-dependent DNA polymerases are highly stereospecific with regard to dNTPs, whereas template-independent terminal deoxynucleotidyl transferase shows less stereodifferentiation. It is likely that the active center of the latter enzyme forms no specific contacts with the nucleic bases of both nucleotide substrate and oligonucleotide primer.
研究了脱氧核糖呋喃糖C-1'和C-4'碳原子的dNTP的所有四种可能的立体异构体作为几种依赖模板的DNA聚合酶和不依赖模板的末端脱氧核苷酸转移酶的底物。结果表明，来自人胎盘的DNA聚合酶α、β和ε以及人免疫缺陷病毒和禽成髓细胞瘤病毒的逆转录酶仅将天然β-D-dNTP掺入DNA链中，而小牛胸腺末端脱氧核苷酸转移酶掺入α-D-dNTP 的两个核苷酸残基，并在β-D-dNTP存在下延伸所得的寡核苷酸。后一种酶在β-D-dNTP存在下也能延伸α-异头D-寡脱氧核苷酸引物。所研究的酶均未使用L-dNTP。这些数据表明，依赖模板的DNA聚合酶对dNTP具有高度的立体特异性，而不依赖模板的末端脱氧核苷酸转移酶的立体分化作用较小。后一种酶的活性中心可能与核苷酸底物和寡核苷酸引物的核酸碱基没有形成特异性接触。
