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Unexpected enzymatic function of an ancient nucleic acid-binding fold.
Rylan R Watkins1, Stella Bockelman1, Anna Vradi1
1Department of Chemistry and Biochemistry, Center for RNA Biology, Ohio State University, Columbus, OH, 43220, United States.
Trypanosoma brucei MCP1, a protein with an OB-fold, unexpectedly deacylates Ala-tRNAs, ensuring translation fidelity. This conserved function highlights an ancient nucleic acid binding domain
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Aminoacyl-tRNA synthetases (ARSs) are crucial for protein synthesis fidelity.
- Eukaryotic ARSs assemble into a multi-aminoacyl-tRNA synthetase complex (MSC).
- The Trypanosoma brucei MSC contains OB-fold proteins MCP1 and MCP2, and the deacylase MCP3.
Purpose of the Study:
- To investigate the unexplored enzymatic activity of MCP1.
- To determine the role of MCP1 within the Trypanosoma brucei MSC.
- To understand the functional significance of OB-fold proteins in tRNA metabolism.
Main Methods:
- Recombinant expression and purification of MCP1.
- In vitro deacylation assays using Ala-tRNAs.
- Domain deletion and site-directed mutagenesis of MCP1.
- Cross-species complementation assays using Saccharomyces cerevisiae Arc1p.
Main Results:
- Recombinant MCP1 exhibits Ala-tRNA deacylation activity.
- The OB-fold of MCP1 contains the catalytic pocket for deacylation.
- Key residues (K326, R331, S335) within the OB-fold are essential for activity.
- MCP1's deacylation function is conserved in Saccharomyces cerevisiae.
Conclusions:
- MCP1 possesses a novel, conserved tRNA deacylation activity.
- The OB-fold domain has an unexpected enzymatic function in nucleic acid metabolism.
- This finding explains the 3' CCA-end binding activity of this protein family.
- The study reveals an ancient function of OB-fold domains in maintaining translational accuracy.
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