High processivity of the reverse transcriptase from a non-long terminal repeat retrotransposon
High processivity of the reverse transcriptase from a non-long terminal repeat retrotransposon
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DOI：
10.1074/jbc.m204345200
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发表时间：
2002-09-20
影响因子：
4.8
通讯作者：
Eickbush, TH
中科院分区：
文献类型：
--
作者：
Bibillo, A;Eickbush, TH
R2是一个反转录转座因子，特异性插入节肢动物的28 S rRNA基因。该元件编码具有内切核酸酶活性和逆转录酶（RT）活性的单一蛋白质，所述内切核酸酶活性切割28 S基因靶位点，所述逆转录酶（RT）活性使用切割的DNA引发逆转录。在这里，我们比较了R2 RT活性的各种特性，以及表征逆转录病毒RT，禽成髓细胞瘤病毒（AMV）。在使用异质RNA模板的持续合成能力测定中，R2 RT可以合成超过AMV RT合成长度的两倍的cDNA，并且在使用poly（rA）模板的测定中可以合成超过AMV RT 4倍的cDNA。与逆转录病毒RT相比，R2 RT的持续合成能力更高，这是由于该酶与RNA模板的解离速率较慢。两种酶的延伸率相似。最后，与逆转录病毒酶相比，R2 RT的一个非常独特的特性是它能够在cDNA合成过程中置换与RNA模板退火的RNA链。我们认为，R2 RT与逆转录病毒RT相比具有更高的持续合成能力和置换特性，这是由于R2蛋白对其活性位点上游的RNA模板具有更大的亲和力。
R2 is a retrotransposable element that specifically inserts into the 28 S rRNA genes of arthropods. The element encodes a single protein with endonuclease activity that cleaves the 28 S gene target site and reverse transcriptase (RT) activity that uses the cleaved DNA to prime reverse transcription. Here we compare various properties of the R2 RT activity with those of the well characterized retroviral RT, avian myeloblastosis virus (AMV). In processivity assays using heterogeneous RNA templates, R2 RT can synthesize cDNA over twice the length of that synthesized by AMV RT and can synthesize cDNA over 4 times longer than AMV RT in assays with poly(rA) templates. The higher processivity of R2 RT compared with retroviral RTs is a result of the slower rate of dissociation of the enzyme from RNA templates. The elongation rates of the two enzymes are similar. Finally, a highly distinct property of the R2 RT, compared with retroviral enzymes, is its ability to displace RNA strands annealed to RNA templates during cDNA synthesis. We suggest that both the higher processivity and displacement properties of R2 RT compared with retroviral RT result from the greater affinity of the R2 protein for the RNA template upstream of its active site.