Identification of Eukaryotic and Prokaryotic Methylthiotransferase for Biosynthesis of 2-Methylthio-N6-threonylcarbamoyladenosine in tRNA
Identification of Eukaryotic and Prokaryotic Methylthiotransferase for Biosynthesis of 2-Methylthio-N6-threonylcarbamoyladenosine in tRNA
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DOI：
10.1074/jbc.m110.106831
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发表时间：
2010-09-10
影响因子：
4.8
通讯作者：
Atta, Mohamed
中科院分区：
文献类型：
--
作者：
Arragain, Simon;Handelman, Samuel K.;Atta, Mohamed
细菌和真核生物的转移RNA都含有高度修饰的腺苷，2-甲硫基-N-6-苏氨酰氨基甲酰腺苷，位于反密码子3‘端附近的37(A(37))位，对于核糖体高效和高精度的蛋白质翻译是必不可少的。利用生物信息学序列分析和活体分析相结合的高效液相色谱/质谱仪技术，我们从不同的序列特征中鉴定出两个亚家族的甲硫基转移酶(MTTase)，分别是细菌细胞中的MtaB和真核细胞和古生物中的e-MtaB。这两个亚家族都负责将N-6-苏氨酰氨基甲酰腺苷转化为2-甲硫基-N-6-苏氨酸氨基甲酰腺苷。最近，人类CDKAL1基因中属于e-MtaB亚家族的一个变异被证明是2型糖尿病的易感基因。因此，CDKAL1是迄今为止发现的第一个真核细胞MTTase。利用CDKAL1细菌同源物枯草芽孢杆菌MtaB(YqeV)的纯化制剂，我们证明了YqeV/CDKAL1酶与先前研究的MTTase MIAB和RIMO一样，含有两个[4Fe-4S]簇。这项工作为阐明CDKAL1的功能奠定了基础。
Bacterial and eukaryotic transfer RNAs have been shown to contain hypermodified adenosine, 2-methylthio-N-6-threonylcarbamoyladenosine, at position 37 (A(37)) adjacent to the 3'-end of the anticodon, which is essential for efficient and highly accurate protein translation by the ribosome. Using a combination of bioinformatic sequence analysis and in vivo assay coupled to HPLC/MS technique, we have identified, from distinct sequence signatures, two methylthiotransferase (MTTase) subfamilies, designated as MtaB in bacterial cells and e-MtaB in eukaryotic and archaeal cells. Both subfamilies are responsible for the transformation of N-6-threonylcarbamoyladenosine into 2-methylthio-N-6-threonylcarbamoyladenosine. Recently, a variant within the human CDKAL1 gene belonging to the e-MtaB subfamily was shown to predispose for type 2 diabetes. CDKAL1 is thus the first eukaryotic MTTase identified so far. Using purified preparations of Bacillus subtilis MtaB (YqeV), a CDKAL1 bacterial homolog, we demonstrate that YqeV/CDKAL1 enzymes, as the previously studied MTTases MiaB and RimO, contain two [4Fe-4S] clusters. This work lays the foundation for elucidating the function of CDKAL1.