Perreault J P, Altman S
Department of Biology, Yale University, New Haven, CT 06520.
J Mol Biol. 1992 Jul 20;226(2):399-409. doi: 10.1016/0022-2836(92)90955-j.
The role of 2'-hydroxyl groups in a model substrate for RNase P from Escherichia coli was studied using mixed DNA/RNA derivatives of such a substrate. The presence of the 2'-hydroxyl groups of nucleotides at positions -1 and -2 in the leader sequence and at position 1, as well as at the first C in the 3'-terminal CCA sequence, are important but not absolutely essential for efficient cleavage of the substrate by RNase P or its catalytic RNA subunit, M1 RNA. The 2'-hydroxyl groups in the substrate that are important for efficient cleavage also participate in the binding of Mg2+. An all-DNA external guide sequence (EGS) can efficiently render a potential substrate, derived from the model substrate, susceptible to cleavage by the enzyme or its catalytic RNA subunit. Furthermore, both DNA and RNA EGSs turn over during the reaction with RNase P in vitro. The identity of the nucleotide at position 1 in the substrate, the adjacent Mg(2+)-binding site in the leader sequence, and the junction of the single and double-stranded regions are the important elements in the recognition of model substrates, as well as in the identification of the sites of cleavage in those model substrates.
利用这种底物的DNA/RNA混合衍生物，研究了2'-羟基在大肠杆菌RNase P模型底物中的作用。前导序列中-1和-2位核苷酸以及1位的2'-羟基的存在，以及3'-末端CCA序列中第一个C的2'-羟基的存在，对于RNase P或其催化RNA亚基M1 RNA有效切割底物很重要，但并非绝对必要。底物中对有效切割很重要的2'-羟基也参与Mg2+的结合。全DNA外部引导序列（EGS）可以有效地使源自模型底物的潜在底物易于被该酶或其催化RNA亚基切割。此外，在体外与RNase P的反应过程中，DNA和RNA EGS都会周转。底物中1位核苷酸的身份、前导序列中相邻的Mg(2+)结合位点以及单链和双链区域的交界处是识别模型底物以及确定这些模型底物中切割位点的重要元素。
