Archaeosine (G) is an archaea-specific tRNA modification synthesized multiple steps. In the first step, archaeosine tRNA guanine transglucosylase (ArcTGT) exchanges the G15 base in tRNA with 7-cyano-7-deazaguanine (preQ). In Euryarchaea, preQ15 in tRNA is further modified by archaeosine synthase (ArcS). ArcS catalyzes a lysine-transfer reaction to produce preQ-lysine (preQ-Lys) as an intermediate. The resulting preQ-Lys15 in tRNA is converted to G15 by a radical S-adenosyl-L-methionine enzyme for archaeosine formation (RaSEA), which forms a complex with ArcS. Here, we focus on the substrate tRNA recognition mechanism of ArcS. Kinetic parameters of ArcS for lysine and tRNA-preQ were determined using a purified enzyme. RNA fragments containing preQ were prepared from tRNA-preQ15. ArcS transferred C-labeled lysine to RNA fragments. Furthermore, ArcS transferred lysine to preQ nucleoside and preQ nucleoside 5′-monophosphate. Thus, the L-shaped structure and the sequence of tRNA are not essential for the lysine-transfer reaction by ArcS. However, the presence of D-arm structure accelerates the lysine-transfer reaction. Because ArcTGT from thermophilic archaea recognizes the common D-arm structure, we expected the combination of ArcTGT and ArcS and RaSEA complex would result in the formation of preQ-Lys15 in all tRNAs. This hypothesis was confirmed using 46 tRNA transcripts and three tRNA transcripts. In addition, ArcTGT did not exchange the preQ-Lys15 in tRNA with guanine or preQ base, showing that formation of tRNA-preQ-Lys by ArcS plays a role in preventing the reverse reaction in G biosynthesis.

中文翻译：

---

来自 Thermococcus kodakarensis 的 ArcS 将 L-赖氨酸转移到 preQ0 核苷衍生物作为最小底物 RNA


古菌素 (G) 是经过多步合成的古菌特异性 tRNA 修饰。第一步，古氨酸 tRNA 鸟嘌呤转葡糖基酶 (ArcTGT) 将 tRNA 中的 G15 碱基与 7-氰基-7-脱氮鸟嘌呤 (preQ) 交换。在广古菌中，tRNA 中的 preQ15 被古氨酸合酶 (ArcS) 进一步修饰。 ArcS 催化赖氨酸转移反应生成 preQ-赖氨酸 (preQ-Lys) 作为中间体。 tRNA 中产生的 preQ-Lys15 通过用于古氨酸形成的自由基 S-腺苷-L-甲硫氨酸酶 (RaSEA) 转化为 G15，与 ArcS 形成复合物。在这里，我们重点关注ArcS的底物tRNA识别机制。使用纯化的酶测定 ArcS 赖氨酸和 tRNA-preQ 的动力学参数。从 tRNA-preQ15 制备含有 preQ 的 RNA 片段。 ArcS 将 C 标记的赖氨酸转移到 RNA 片段上。此外，ArcS 将赖氨酸转移至 preQ 核苷和 preQ 核苷 5'-单磷酸。因此，L 形结构和 tRNA 序列对于 ArcS 的赖氨酸转移反应不是必需的。然而，D 臂结构的存在加速了赖氨酸转移反应。由于来自嗜热古菌的 ArcTGT 识别常见的 D 臂结构，因此我们预计 ArcTGT 与 ArcS 和 RaSEA 复合物的组合将导致所有 tRNA 中 preQ-Lys15 的形成。使用 46 个 tRNA 转录本和 3 个 tRNA 转录本证实了这一假设。此外，ArcTGT没有将tRNA中的preQ-Lys15与鸟嘌呤或preQ碱基进行交换，表明ArcS形成的tRNA-preQ-Lys起到了防止G生物合成逆反应的作用。