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Updated: May 20, 2026
An In Vitro Assay to Detect tRNA-Isopentenyl Transferase Activity
Published on: October 8, 2018
Structural basis for the activity of a cytoplasmic RNA terminal uridylyl transferase
Luke A Yates1, Sophie Fleurdépine2, Olivia S Rissland2
1Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, UK.
Abstract:
Cytoplasmic terminal uridylyl transferases comprise a conserved family of enzymes that negatively regulate the stability or biological activity of a variety of eukaryotic RNAs, including mRNAs and tumor-suppressor let-7 microRNAs. Here we describe crystal structures of the Schizosaccharomyces pombe cytoplasmic terminal uridylyl transferase Cid1 in two apo conformers and bound to UTP. We demonstrate that a single histidine residue, conserved in mammalian Cid1 orthologs, is responsible for discrimination between UTP and ATP. We also describe a new high-affinity RNA substrate-binding mechanism of Cid1, which is essential for enzymatic activity and is mediated by three basic patches across the surface of the enzyme. Overall, our structures provide a basis for understanding the activity of Cid1 and a mechanism of UTP selectivity conserved in its human orthologs, suggesting potential implications for anticancer drug design.
Insights
Cytoplasmic terminal uridylyl transferases like Cid1 regulate RNA stability. Structural studies reveal Cid1
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Cytoplasmic terminal uridylyl transferases (TUTases) are enzymes regulating RNA stability and activity.
- These enzymes impact messenger RNAs (mRNAs) and microRNAs (miRNAs), including tumor-suppressor let-7 miRNAs.
Purpose of the Study:
- To elucidate the structural basis of Cid1 enzyme activity and substrate specificity.
- To understand the mechanism of UTP selectivity in Cid1 and its human orthologs.
Main Methods:
- X-ray crystallography was used to determine the structures of Schizosaccharomyces pombe Cid1.
- Structures were obtained in apo conformations and when bound to UTP (uridine triphosphate).
Main Results:
- Crystal structures revealed two apo conformers and a UTP-bound form of Cid1.
- A conserved histidine residue dictates UTP versus ATP discrimination.
- A novel high-affinity RNA substrate-binding mechanism involving three basic patches was identified.
Conclusions:
- The determined structures provide insights into Cid1 function and UTP selectivity.
- The conserved UTP selectivity mechanism suggests potential applications in anticancer drug design.
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