Dataset for: The epsilon motif of Hepatitis B virus RNA exhibits a potassium dependent ribonucleolytic activity

Fourteen different classes of ribozymes are known that catalyze a range of diverse chemical reactions. We report here a novel potassium-dependent nucleolytic activity present in Hepatitis B Virus (HBV) RNA. A short RNA region (53 nt) with enzymatic properties released itself from the viral sequence by cis cleavages and could subsequently act in trans. The released region encompassed the epsilon motif present in the HBV RNA. The 3’ end of the liberated fragment was within the non-canonical polyadenylation signal (UAUAAA) of the viral RNA while cleavages at about 53 nt upstream sites released the fragment. Mutations of the primary scissile sites or annealing these sites to the antisense oligodeoxyribonucleotides blocked the release of this short fragment and annulled subsequent trans cleavage activity. An exogenously synthesized short transcript of only this 53 nt, was active as a sequence independent trans-acting nuclease and cleaved after pyrimidines in viral or other substrate transcripts under physiological potassium ion concentrations. Formation of a G-quadruplex within this region was suggested by circular dichroism and nondenaturing polyacrylamide gel analyses. Our results reveal a unique natural example of a trans acting ribonuclease that cleaves at multiple sites in a sequence independent fashion. The presence of this novel activity implores a dynamic structural behaviour in the epsilon region and raises new questions about HBV gene regulation.

目前已知有十四类不同的核酶（ribozyme）可催化一系列多样化的化学反应。本研究报道了一种存在于乙型肝炎病毒（Hepatitis B Virus, HBV）RNA中的新型钾离子依赖型核酸水解活性。一段具备酶学特性的53个核苷酸（nucleotide, nt）的RNA区域可通过顺式切割（cis cleavage）从病毒序列中自我释放，并可后续以反式（trans）模式发挥功能。该释放出的RNA区域包含了HBV RNA中存在的ε基序（epsilon motif）。该释放片段的3’端位于病毒RNA的非经典多聚腺苷酸化信号（polyadenylation signal, UAUAAA）区域内，而在其上游约53 nt位点处发生的切割则可释放该片段。对主要切割位点（scissile site）进行突变，或将这些位点与反义寡脱氧核糖核苷酸（oligodeoxyribonucleotide）进行退火结合，均可阻断该短片段的释放，并消除后续的反式切割活性。仅由该53 nt区域构成的外源合成短转录本，可作为不依赖序列的反式作用核酸酶（trans-acting nuclease）发挥活性；在生理钾离子浓度条件下，该转录本可于病毒或其他底物转录本的嘧啶（pyrimidine）位点下游进行切割。圆二色谱（circular dichroism）与非变性聚丙烯酰胺凝胶（nondenaturing polyacrylamide gel）分析结果表明，该区域内可形成G-四链体（G-quadruplex）。本研究结果揭示了一种独特的天然反式作用核糖核酸酶（ribonuclease）实例，该酶可不以序列依赖的方式在多个位点进行切割。这种新型活性的存在，提示ε基序区域具有动态结构特性，并为HBV基因调控领域提出了新的科学问题。