Pongracz K, Dosanjh M K, Singer B, Bodell W J
Department of Neurological Surgery, University of California San Francisco, School of Medicine 94143-0806.
Carcinogenesis. 1994 Jul;15(7):1371-5. doi: 10.1093/carcin/15.7.1371.
A diastereomeric mixture of the regioisomers O6-(2-hydroxy-2-phenylethyl)-2'-deoxyguanosine (st6G, beta-isomer) and O6-(2-hydroxy-1-phenylethyl)-2'-deoxyguanosine (alpha-isomer) was site-specifically placed in a 25 base oligonucleotide template 5'-CCGCTAst6GCGGGTACCGAGCTCGAAT-3' using CED phosphoramidite chemistry. Using 32P-post-labeling we found the oligonucleotide to contain 95% of the beta-isomer and 5% of the alpha-isomer of st6G. st6G as the 3'-phosphate was found to be considerably more acid labile than O6-methyl-2'-deoxyguanosine-3'-phosphate, leading to dealkylation during oligonucleotide synthesis. The diastereomeric mixture of O6-(2-hydroxy-2-phenylethyl)-2'-deoxy-guanosine-5'-triphosphate (st6dGTP) was chemically synthesized and used as a substrate for the exonuclease-free Klenow fragment of Escherichia coli DNA polymerase I. This study demonstrated that st6dGTP could be incorporated opposite deoxycytidine and did not completely block replication.
区域异构体O6-(2-羟基-2-苯乙基)-2'-脱氧鸟苷（st6G，β-异构体）和O6-(2-羟基-1-苯乙基)-2'-脱氧鸟苷（α-异构体）的非对映异构体混合物通过CED亚磷酰胺化学方法位点特异性地置于25个碱基的寡核苷酸模板5'-CCGCTAst6GCGGGTACCGAGCTCGAAT-3'中。使用32P后标记，我们发现该寡核苷酸含有95%的st6G的β-异构体和5%的α-异构体。发现作为3'-磷酸酯的st6G比O6-甲基-2'-脱氧鸟苷-3'-磷酸酯对酸更不稳定，导致在寡核苷酸合成过程中发生脱烷基化。化学合成了O6-(2-羟基-2-苯乙基)-2'-脱氧鸟苷-5'-三磷酸酯（st6dGTP）的非对映异构体混合物，并将其用作大肠杆菌DNA聚合酶I无外切核酸酶的Klenow片段的底物。这项研究表明，st6dGTP可以与脱氧胞苷配对掺入，并且不会完全阻断复制。
