Systematic substitution of individual bases in two important single‐stranded regions of the HDV ribozyme for evaluation of the role of specific bases

FEBS Letters

Suh, Young-Ah¹  Taira, Kazunari¹  Kawakami, Junji¹  Nishikawa, Fumiko¹  Kumar, P.K.R.¹  Nishikawa, Satoshi¹ 

[1] National Institute of Bioscience and Human-Technology, Agency of Industrial Science and Technology, MITI, Tsukuba Science City, 305 Japan

关键词: HDV ribozyme;    Self-cleavage activity;    Pseudoknot structure;    Single-stranded region;    Point mutation;    HDV;    hepatitis delta virus;    PEG;    polyethylene glycol;    BPB;    Bromophenol blue;    XCFF;    xylene cyanol FF;   

DOI  :  10.1016/0014-5793(93)81782-U

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

To elucidate the role of specific bases in the self-cleavage activity of the human hepatitis delta virus (HDV) ribozyme, systematic substitutions of individual bases in two important single-stranded regions [between nucleotides 726–731 (SSrA region) and 762–766 (SSrB region)] were carried out by oligonucleotide-directed point mutagenesis. Among the mutants obtained, 12 mutants (G726 variants, G727A, G727C, G728C, G762A, G762C, C763 variants and A766C) could not tolerate the respective base-substitutions and self-cleavage activities were reduced to very low levels (10%), suggesting a requirement of the respective bases. In particular, G726 in the SSrA region and C763 in the SSrB region were found to be essential for the ribozyme activity. We could determine the preferred sequences, 5′-G-G-(G/A/U)-N-(A/U/G)-Pu-3′for SSrA and 5′-(G/U)-C-N-(A/G/U)-A-3′ for SSrB regions, respectively.

【 授权许可】

Unknown