Department of Biochemistry, Duke University School of Medicine, Durham, North Carolina 27710, USA.
RNA. 2020 Jan;26(1):69-82. doi: 10.1261/rna.072231.119. Epub 2019 Nov 8.
Most mitochondrial mRNAs are transcribed as polycistronic precursors that are cleaved by endonucleases to produce mature mRNA transcripts. However, recent studies have shown that mitochondrial transcripts in the kinetoplastid protozoan, , are transcribed individually. Also unlike most mitochondrial mRNAs, the 5' end of these transcripts harbor a triphosphate that is hydrolyzed. This modification is carried out by a putative Nudix hydrolase called MERS1. The Nudix motif in MERS1 is degenerate, lacking a conserved glutamic acid, thus it is unclear how it may bind its substrates and whether it contains a Nudix fold. To obtain insight into this unusual hydrolase, we determined structures of apo, GTP-bound and RNA-bound MERS1 to 2.30 Å, 2.45 Å, and 2.60 Å, respectively. The MERS1 structure has a unique fold that indeed contains a Nudix motif. The nucleotide bound structures combined with binding studies reveal that MERS1 shows preference for RNA sequences with a central guanine repeat which it binds in a single-stranded conformation. The apo MERS1 structure indicates that a significant portion of its nucleotide binding site folds upon substrate binding. Finally, a potential interaction region for a binding partner, MERS2, that activates MERS1 was identified. The MERS2-like peptide inserts a glutamate near the missing Nudix acidic residue in the RNA binding pocket, suggesting how the enzyme may be activated. Thus, the combined studies reveal insight into the structure and enzyme properties of MERS1 and its substrate-binding activities.
大多数线粒体 mRNA 是作为多顺反子前体转录的，这些前体通过内切酶切割产生成熟的 mRNA 转录本。然而，最近的研究表明，动基体原生动物中的线粒体转录物是单独转录的。与大多数线粒体 mRNA 不同，这些转录物的 5' 端含有一个被水解的三磷酸基团。这种修饰是由一种称为 MERS1 的假定 Nudix 水解酶完成的。MERS1 中的 Nudix 基序是退化的，缺乏保守的谷氨酸，因此不清楚它如何结合其底物，以及它是否含有 Nudix 折叠。为了深入了解这种不寻常的水解酶，我们分别确定了 apo、GTP 结合和 RNA 结合的 MERS1 结构，分辨率分别为 2.30Å、2.45Å 和 2.60Å。MERS1 结构具有独特的折叠，确实含有 Nudix 基序。结合结合研究的核苷酸结合结构表明，MERS1 优先结合具有中央鸟嘌呤重复序列的 RNA 序列，它以单链构象结合。apo MERS1 结构表明，其核苷酸结合位点的很大一部分在底物结合时折叠。最后，确定了一个与激活 MERS1 的结合伙伴 MERS2 的潜在相互作用区域。MERS2 样肽在 RNA 结合口袋中插入一个谷氨酸，靠近缺失的 Nudix 酸性残基，这表明该酶可能被激活。因此，综合研究揭示了 MERS1 及其底物结合活性的结构和酶特性的深入了解。
