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首页> 外文期刊>Biochemistry >The Human HDV-like CPEB3 Ribozyme Is Intrinsically Fast-Reacting
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The Human HDV-like CPEB3 Ribozyme Is Intrinsically Fast-Reacting

机译：人类HDV样CPEB3核酶本质上是快速反应的。

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摘要

Self-cleaving RNAs have recently been identified in mammalian genomes. A small ribozyme related in structure to the hepatitis delta virus (H DV) ribozyme occurs in a number of mammals, including chimpanzees and humans, within an intron of the CPEB3 gene. The catalytic mechanisms for the CPEB3 and HDV ribozymes appear to be similar, generating cleavage products with 5'-hydroxyl and 2',3'-cyclic phosphate termini; nonetheless, the cleavage rate reported for the CPEB3 ribozyme is more than 6000-fold slower than for the fastest HDV ribozyme. Herein, we use full-length RNA and cotranscriptional self-cleavage assays to compare reaction rates among human CPEB3, chimp CPEB3, and H DV ribozymes. Our data reveal that a single base change of the upstream flanking sequence, which sequesters an intrinsically weak P1.1 pairing in a misfold, increases the rate of the wild-type human CPEB3 ribozyme by similar to 250-fold; thus, the human ribozyme is intrinsically fast-reacting. Secondary structure determination and native gel analyses reveal that the cleaved population of the CPEB3 ribozyme has a single, secondary structure that closely resembles the HDV ribozyme. In contrast, the precleavage population of the CPEB3 ribozyme appears to have a more diverse secondary structure, possibly reflecting misfolding with the upstream sequence and dynamics intrinsic to the ribozyme. Prior identification of expressed sequence tags (ESTs) in human cells indicated that cleavage activity of the human ribozyme is tissue-specific. It is therefore possible that cellular factors interact with regions upstream of the CPEB3 ribozyme to unmask its high intrinsic reactivity.
机译：最近已经在哺乳动物基因组中鉴定了自切割RNA。在结构上与肝炎三角洲病毒（H DV）核酶有关的一种小的核酶，存在于CPEB3基因内含子内的许多哺乳动物中，包括黑猩猩和人类。 CPEB3和HDV核酶的催化机制似乎相似，产生具有5'-羟基和2'，3'-环状磷酸酯末端的裂解产物。尽管如此，据报道，CPEB3核酶的裂解速率比最快的HDV核酶慢6000倍以上。在本文中，我们使用全长RNA和共转录自我切割测定法来比较人CPEB3，黑猩猩CPEB3和H DV核酶之间的反应速率。我们的数据显示，上游侧翼序列的一个碱基改变，将一个本来就很弱的P1.1配对以错误折叠的方式螯合，可使野生型人CPEB3核酶的发生率增加约250倍。因此，人类核酶本质上是快速反应的。二级结构测定和天然凝胶分析表明，CPEB3核酶的裂解种群具有单个类似于HDV核酶的二级结构。相反，CPEB3核酶的预切割种群似乎具有更多样化的二级结构，可能反映了上游序列的错误折叠和核酶固有的动力学。事先鉴定人细胞中表达的序列标签（EST）表明，人核酶的切割活性是组织特异性的。因此，细胞因子可能会与CPEB3核酶上游区域相互作用，从而掩盖其高固有反应性。

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