Two reactions of Haloferax voicanii RNA splicing enzymes:: Joining of exons and circularization of introns
Two reactions of Haloferax voicanii RNA splicing enzymes:: Joining of exons and circularization of introns
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作者：
Salgia, SR;Singh, SK;Gupta, R
RNA剪接涉及至少两种蛋白酶，一种特异性核酸内切酶和一种特异性连接酶。核酸内切酶识别并在特征性凸起-螺旋-凸起（BHB）结构内切割，该结构通过两个外显子-内含子连接附近的区域配对形成，产生2 '，3'-环磷酸和5 '-羟基末端。连接酶连接外显子并将环状磷酸转化为连接磷酸。连接产物含有七碱基发夹环，其中剪接点位于环的两个3'末端残基之间。内切剪接酶也参与rRNA加工，在通过rRNA的5'和3'侧翼区配对形成的BHB结构内切割。来自前体rRNA的大的游离内含子在某些泉古菌中被观察到是稳定和丰富的环状RNA。在本研究中，我们表明，Haloferax volcano ii，一个Euryarchaeote的细胞，含有环状RNA的3 '，5'-磷酸二酯连接之间的内含子的两个末端来自其前tRNA。H. volcanii连接酶在体外也可以环化内切核酸酶切割的内含子和非内切核酸酶产生的底物。外显子连接和内含子环化是可以独立发生的机制相似的连接反应。在体外连接反应中，连接的发夹环的大小和该环内剪接点的位置可以改变。总的来说，古细菌RNA剪接似乎涉及两组对称的酯交换反应。
Archaeal RNA splicing involves at least two protein enzymes, a specific endonuclease and a specific ligase. The endonuclease recognizes and cleaves within a characteristic bulge-helix-bulge (BHB) structure formed by pairing of the regions near the two exon-intron junctions, producing 2',3'-cyclic phosphate and 5'-hydroxyl termini. The ligase joins the exons and converts the cyclic phosphate into junction phosphate. The ligated product contains a seven-base hairpin loop, in which the splice junction is in between the two 3' terminal residues of the loop. Archaeal splicing endonucleases are also involved in rRNA processing, cutting within the BHB structures formed by pairing of the 5' and 3' flanking regions of the rRNAs. Large free introns derived from pre-rRNAs have been observed as stable and abundant circular RNAs in certain Crenarchaeota, a kingdom in the domain Archaea. In the present study, we show that the cells of Haloferax volcanii, a Euryarchaeote, contain circular RNAs formed by 3',5'-phosphodiester linkage between the two termini of the introns derived from their pre-tRNAs. H. volcanii ligase, in vitro, can also circularize both endonuclease-cleaved introns, and non-endonuclease-produced substrates. Exon joining and intron circularization are mechanistically similar ligation reactions that can occur independently. The size of the ligated hairpin loop and position of the splice junction within this loop can be changed in in vitro ligation reactions. Overall, archaeal RNA splicing seems to involve two sets of two symmetric transesterification reactions each.