Constructing an efficient trans‐acting genomic HDV ribozyme

FEBS Letters

Yuda, Kazuhiro¹  Suh, Young-Ah¹  Taira, Kazunari¹  Kawakami, Junji²  Nishikawa, Fumiko¹  Kumar, Penmetcha K.R.¹  Ohtsuka, Eiko²  Maeda, Hidekatsu¹  Nishikawa, Satoshi¹ 

[1] National Institute of Bioscience and Human Technology, Agency of Industrial Science and Technology, MITI, Tsukuba Science City 305, Japan;Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060, Japan

关键词: HDV ribozyme;    Pseudoknot structure;    Ribozyme;    trans-acting;   

DOI  :  10.1016/0014-5793(96)00941-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We have engineered a genomic HDV ribozyme to construct several trans-acting ribozymes for use in trans to cleave target RNAs. Among the 10 different combinations attempted, only HDV88-Trans had cleavage activity on the 13-nucleotide substrate, R13, in vitro. To improve the cleavage efficiency, at least in vitro, of the HDV88-Trans ribozyme (k_clv = 0.022 min⁻¹), we have constructed several variants that differ in forming stem II (length) in the pseudoknot secondary structure model. When cleavage rate constants were analyzed and compared among variants of HDV88-Trans, HDV88-Trans-4 yielded k_clv = 1.7 min⁻¹. HDV88-Trans-4 thus represents the highest active genomic HDV ribozyme that functions in trans thus far constructed, and has activity under physiological conditions (pH 7.1 at 37°C with 1 mM of MgCl₂).

【 授权许可】

Unknown