Targeting sites within HIV-1 cDNA with a DNA-cleaving ribozyme.
Targeting sites within HIV-1 cDNA with a DNA-cleaving ribozyme.
复制标题
使用 DNA 切割核酶靶向 HIV-1 cDNA 内的位点。
DOI：
10.1021/bi960845g
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Joyce,GF
中科院分区：
文献类型：
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作者：
Raillard,SA;Joyce,GF
在模拟生理条件下有效切割单链DNA的四膜虫核酶的变体[Tsang，J.，& Joyce，G. F.（1994）Biochemistry 33，5966 - 5973]基于其切割对应于HIV-1 cDNA内的保守靶位点的合成寡核苷酸底物的能力而被评价为潜在的治疗剂。为了增加核酶的序列选择性，其底物识别结构域从6个核苷酸延伸到12个核苷酸，允许与位于切割位点上游和下游的底物核苷酸进行碱基配对。可以改变延伸的识别结构域的序列以允许切割多种不同的DNA靶标。该核酶具有高度的序列特异性，对与核酶识别结构域形成单碱基错配的底物的识别能力可达102 ~ 104倍。发生在切割位点附近的错配与发生在较远位置的错配相比，导致活性降低更大。
A variant of theTetrahymenaribozyme that efficiently cleaves single-stranded DNA under simulated physiological conditions [Tsang, J., & Joyce, G. F. (1994)Biochemistry 33, 5966−5973] was evaluated as a potential therapeutic agent on the basis of its ability to cleave synthetic oligonucleotide substrates corresponding to conserved target sites within HIV-1 cDNA. In order to increase the sequence selectivity of the ribozyme, its substrate recognition domain was extended from 6 to 12 nucleotides, allowing base pairing with substrate nucleotides that lie both upstream and downstream of the cleavage site. The sequence of the extended recognition domain could be changed to allow cleavage of a variety of different DNA targets. The ribozyme exhibited a high degree of sequence specificity, discriminating by a factor of 102to more than 104against substrates that form a single-base mismatch with the ribozyme's recognition domain. Mismatches that occurred close to the cleavage site led to a greater decrease in activity compared to those that occurred farther away.