Allemand Frédéric, Mathy Nathalie, Brechemier-Baey Dominique, Condon Ciarán
UPR 9073, Université de Paris 7-Denis Diderot, Institut de Biologie Physico-Chimique 13 rue Pierre et Marie Curie, 75005 Paris, France.
Nucleic Acids Res. 2005 Aug 2;33(13):4368-76. doi: 10.1093/nar/gki752. Print 2005.
The maturation of 5S ribosomal RNA in low G+C Gram-positive bacteria is catalyzed by a highly conserved, approximately 190 residue, enzyme, called ribonuclease M5 (RNase M5). Sequence alignment had predicted that the N-terminal half of RNase M5 would consist of a Toprim domain, a protein fold found in type IA and type II topoisomerases, DnaG-like primases, OLD family nucleases and RecR proteins [L. Aravind, D. D. Leipe and E. V. Koonin (1998) Nucleic Acids Res., 26, 4205-4213]. Here, we present structural modelling data and a mutational analysis of RNase M5 that confirms this hypothesis. The N-terminal half of RNase M5 can be fitted to the Toprim domain of the DnaG catalytic core. Mutation of amino acid residues highly conserved among RNase M5 enzymes and members of the Toprim domain family showed that alteration of residues critical for topoisomerase and primase activity also had a dramatic effect on the cleavage of 5S rRNA precursor by RNase M5 both in vivo and in vitro. This suggests that the mechanisms of double-stranded RNA cleavage by RNase M5 and double-stranded DNA cleavage by members of the topoisomerase family are related.
低G+C含量革兰氏阳性菌中5S核糖体RNA的成熟由一种高度保守、约190个残基的酶催化，该酶称为核糖核酸酶M5（RNase M5）。序列比对预测，RNase M5的N端一半将由一个Toprim结构域组成，这是一种在IA型和II型拓扑异构酶、DnaG样引发酶、OLD家族核酸酶和RecR蛋白中发现的蛋白质折叠结构【L. Aravind、D. D. Leipe和E. V. Koonin（1998年），《核酸研究》，26，4205 - 4213】。在此，我们展示了结构建模数据以及对RNase M5的突变分析，证实了这一假设。RNase M5的N端一半可以适配到DnaG催化核心的Toprim结构域。在RNase M5酶和Toprim结构域家族成员中高度保守的氨基酸残基发生突变表明，对拓扑异构酶和引发酶活性至关重要的残基的改变，在体内和体外对RNase M5切割5S rRNA前体也有显著影响。这表明RNase M5切割双链RNA的机制与拓扑异构酶家族成员切割双链DNA的机制有关。
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