EFFICIENT TRANS-CLEAVAGE OF A STEM-LOOP RNA SUBSTRATE BY A RIBOZYME DERIVED FROM NEUROSPORA VS RNA
EFFICIENT TRANS-CLEAVAGE OF A STEM-LOOP RNA SUBSTRATE BY A RIBOZYME DERIVED FROM NEUROSPORA VS RNA
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DOI：
10.1002/j.1460-2075.1995.tb07011.x
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发表时间：
1995-01-16
期刊：
影响因子：
11.4
通讯作者：
COLLINS, RA
中科院分区：
文献类型：
--
作者：
GUO, HCT;COLLINS, RA
我们构建了一种含有144个核苷酸的Neurospora VS RNA的核酶，可以以真正的酶促方式催化分离RNA的裂解（k - M接近0.13 μ M, k（cat）接近0.7/min）。顺式和反式裂解速率的比较，以及pH对裂解速率的影响，表明在裂解之前发生了一个限速步骤，可能是构象变化。切割所需的最小连续底物序列由切割位点上游1个核苷酸和下游19个核苷酸组成。与大多数其他核酶与其底物的长单链区域相互作用不同，VS核酶的最小底物主要由稳定的茎环组成，这似乎排除了通过广泛的沃森-克里克碱基配对来识别它。
We have constructed a ribozyme containing 144 nucleotides of Neurospora VS RNA that can catalyze the cleavage of a separate RNA in a true enzymatic manner (K-m approximate to 0.13 mu M, k(cat) approximate to 0.7/min). Comparison of the rates of cis- and trans-cleavage, as well as the lack of effect of pH on the rate of cleavage, suggest that a rate-limiting step, possibly a conformational change, occurs prior to cleavage. The minimum contiguous substrate sequence required for cleavage consists of one nucleotide upstream and 19 nucleotides downstream of the cleavage site. Unlike most other ribozymes which interact with long single-stranded regions of their substrates, the minimal substrate for the VS ribozyme consists mostly of a stable stem-loop, which would appear to preclude its recognition simply via extensive Watson-Crick base pairing.