Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306, USA.
RNA. 2011 Feb;17(2):244-50. doi: 10.1261/rna.2415811. Epub 2010 Dec 13.
Cbf5 is the catalytic subunit of the H/ACA small nucleolar/Cajal body ribonucleoprotein particles (RNPs) responsible for site specific isomerization of uridine in ribosomal and small nuclear RNA. Recent evidence from studies on archaeal Cbf5 suggests its second functional role in modifying tRNA U55 independent of guide RNA. In order to act both as a stand-alone and a RNP pseudouridine synthase, Cbf5 must differentiate features in H/ACA RNA from those in tRNA or rRNA. Most H/ACA RNAs contain a hallmark ACA trinucleotide downstream of the H/ACA motif. Here we challenged an archaeal Cbf5 (in the form of a ternary complex with its accessory proteins Nop10 and Gar1) with T-stem-loop RNAs with or without ACA trinucleotide in the stem. Although these substrates were previously shown to be substrates for the bacterial stand-alone pseudouridine synthase TruB, the Cbf5-Nop10-Gar1 complex was only able to modify those without ACA trinucleotide. A crystal structure of Cbf5-Nop10-Gar1 trimer bound with an ACA-containing T-stem-loop revealed that the ACA trinucleotide detracted Cbf5 from the stand-alone binding mode, thereby suggesting that the H/ACA RNP-associated function of Cbf5 likely supersedes its stand-alone function.
Cbf5 是 H/ACA 小核仁/Cajal 体核糖核蛋白颗粒（RNP）的催化亚基，负责核糖体和小核 RNA 中尿嘧啶的特异性异构化。最近对古细菌 Cbf5 的研究证据表明，它在独立于向导 RNA 的情况下修饰 tRNA U55 方面具有第二个功能作用。为了既能作为独立的 RNP 假尿嘧啶合酶发挥作用，Cbf5 必须区分 H/ACA RNA 与 tRNA 或 rRNA 的特征。大多数 H/ACA RNA 在 H/ACA 基序下游含有一个标志性的 ACA 三核苷酸。在这里，我们用含有或不含有茎部 ACA 三核苷酸的 T 茎环 RNA 来挑战一种古细菌 Cbf5（以与其辅助蛋白 Nop10 和 Gar1 的三元复合物的形式）。尽管这些底物以前被证明是细菌独立假尿嘧啶合酶 TruB 的底物，但 Cbf5-Nop10-Gar1 复合物只能修饰那些没有 ACA 三核苷酸的底物。与含有 ACA 的 T 茎环结合的 Cbf5-Nop10-Gar1 三聚体的晶体结构表明，ACA 三核苷酸使 Cbf5 无法采用独立结合模式，这表明 Cbf5 的 H/ACA RNP 相关功能可能取代了其独立功能。
