Department of Biochemistry, Gene Center, Ludwig-Maximilians University Munich, University of Munich, Munich, Germany.
PLoS Biol. 2023 Apr 20;21(4):e3001995. doi: 10.1371/journal.pbio.3001995. eCollection 2023 Apr.
Cotranslational modification of the nascent polypeptide chain is one of the first events during the birth of a new protein. In eukaryotes, methionine aminopeptidases (MetAPs) cleave off the starter methionine, whereas N-acetyl-transferases (NATs) catalyze N-terminal acetylation. MetAPs and NATs compete with other cotranslationally acting chaperones, such as ribosome-associated complex (RAC), protein targeting and translocation factors (SRP and Sec61) for binding sites at the ribosomal tunnel exit. Yet, whereas well-resolved structures for ribosome-bound RAC, SRP and Sec61, are available, structural information on the mode of ribosome interaction of eukaryotic MetAPs or of the five cotranslationally active NATs is only available for NatA. Here, we present cryo-EM structures of yeast Map1 and NatB bound to ribosome-nascent chain complexes. Map1 is mainly associated with the dynamic rRNA expansion segment ES27a, thereby kept at an ideal position below the tunnel exit to act on the emerging substrate nascent chain. For NatB, we observe two copies of the NatB complex. NatB-1 binds directly below the tunnel exit, again involving ES27a, and NatB-2 is located below the second universal adapter site (eL31 and uL22). The binding mode of the two NatB complexes on the ribosome differs but overlaps with that of NatA and Map1, implying that NatB binds exclusively to the tunnel exit. We further observe that ES27a adopts distinct conformations when bound to NatA, NatB, or Map1, together suggesting a contribution to the coordination of a sequential activity of these factors on the emerging nascent chain at the ribosomal exit tunnel.
新生多肽链的共翻译修饰是新蛋白质产生过程中的第一个事件之一。在真核生物中，甲硫氨酰肽酶（MetAPs）切除起始甲硫氨酸，而 N-乙酰转移酶（NATs）催化 N 端乙酰化。MetAPs 和 NATs 与其他共翻译作用的伴侣竞争，如核糖体相关复合物（RAC）、蛋白质靶向和易位因子（SRP 和 Sec61），以争夺核糖体隧道出口的结合位点。然而，尽管有分辨率较高的核糖体结合 RAC、SRP 和 Sec61 的结构，但是关于真核 MetAPs 或五种共翻译活性 NATs 的核糖体相互作用模式的结构信息仅适用于 NatA。在这里，我们展示了酵母 Map1 和 NatB 与核糖体-新生链复合物结合的冷冻电镜结构。Map1 主要与动态 rRNA 扩展片段 ES27a 相关联，从而保持在隧道出口下方的理想位置，以作用于新出现的底物新生链。对于 NatB，我们观察到两个 NatB 复合物的副本。NatB-1 直接结合在隧道出口下方，再次涉及 ES27a，而 NatB-2 位于第二个通用接头位点（eL31 和 uL22）下方。两个 NatB 复合物在核糖体上的结合模式不同，但与 NatA 和 Map1 重叠，这意味着 NatB 仅结合隧道出口。我们进一步观察到，当 ES27a 与 NatA、NatB 或 Map1 结合时，ES27a 采用不同的构象，这共同表明 ES27a 对这些因子在核糖体出口隧道中对新生链的顺序活性的协调有贡献。
