拟南芥中假尿苷-5'-单磷酸糖基化酶 PUMY 的结构和功能。

IF 3.6 3区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY RNA Biology Pub Date : 2024-01-01 Epub Date: 2023-12-20 DOI:10.1080/15476286.2023.2293340
Jeongyun Lee, Sang-Hoon Kim, Sangkee Rhee
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引用次数: 0

摘要

伪尿嘧啶是一种非典型的 C 核苷,含有尿嘧啶和核糖之间的 C-C 糖苷键。在假尿苷的两步降解过程中,假尿嘧啶-5'-单磷酸糖基化酶(PUMY)负责第二步,催化假尿嘧啶-5'-单磷酸(ΨMP)中的 C-C 糖苷键裂解为尿苷和核糖-5'-磷酸,并通过其他代谢途径循环利用。已有关于大肠杆菌 PUMY 结构特征的报道,但ΨMP 底物特异性的细节尚不清楚。在这里,我们展示了拟南芥 PUMY 在不同连接状态下的三个晶体结构以及ΨMP 降解的动力学分析。结果表明,在 PUMY 家族中保守的 Thr149 和 Asn308 是识别 ΨMP 核碱基的结构决定因素。ΨMP与核糖-5'-磷酸的不同结合模式也表明,ΨMP的核碱基而不是磷酸基决定了底物的结合模式。活性位点从开放到封闭的转变是催化所必需的,这种转变是由活性位点附近的两个α-螺旋(α11 和 α12)介导的。突变分析验证了活性位点残基在催化作用中的作用。我们的结构和功能分析进一步揭示了 PUMY 对 ΨMP 的酶学特征。
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Structure and function of the pseudouridine 5'-monophosphate glycosylase PUMY from Arabidopsis thaliana.

Pseudouridine is a noncanonical C-nucleoside containing a C-C glycosidic linkage between uracil and ribose. In the two-step degradation of pseudouridine, pseudouridine 5'-monophosphate glycosylase (PUMY) is responsible for the second step and catalyses the cleavage of the C-C glycosidic bond in pseudouridine 5'-monophosphate (ΨMP) into uridine and ribose 5'-phosphate, which are recycled via other metabolic pathways. Structural features of Escherichia coli PUMY have been reported, but the details of the substrate specificity of ΨMP were unknown. Here, we present three crystal structures of Arabidopsis thaliana PUMY in different ligation states and a kinetic analysis of ΨMP degradation. The results indicate that Thr149 and Asn308, which are conserved in the PUMY family, are structural determinants for recognizing the nucleobase of ΨMP. The distinct binding modes of ΨMP and ribose 5'-phosphate also suggest that the nucleobase, rather than the phosphate group, of ΨMP dictates the substrate-binding mode. An open-to-close transition of the active site is essential for catalysis, which is mediated by two α-helices, α11 and α12, near the active site. Mutational analysis validates the proposed roles of the active site residues in catalysis. Our structural and functional analyses provide further insight into the enzymatic features of PUMY towards ΨMP.

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来源期刊
RNA Biology
RNA Biology 生物-生化与分子生物学
CiteScore
8.60
自引率
0.00%
发文量
82
审稿时长
1 months
期刊介绍: RNA has played a central role in all cellular processes since the beginning of life: decoding the genome, regulating gene expression, mediating molecular interactions, catalyzing chemical reactions. RNA Biology, as a leading journal in the field, provides a platform for presenting and discussing cutting-edge RNA research. RNA Biology brings together a multidisciplinary community of scientists working in the areas of: Transcription and splicing Post-transcriptional regulation of gene expression Non-coding RNAs RNA localization Translation and catalysis by RNA Structural biology Bioinformatics RNA in disease and therapy
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