Abstract
Ribosome biogenesis is a complicated, multistage process coordinated by ribosome assembly factors. Ribosome binding factor A (RbfA) is a bacterial one, which possesses a single structural type-II KH domain. By this domain, RbfA binds to a 16S rRNA precursor in small ribosomal subunits to promote its 5′-end processing. The human RbfA homolog, mtRbfA, binds to 12S rRNAs in the mitoribosomal small subunits and promotes its critical maturation process, the dimethylation of two highly conserved consecutive adenines, which differs from that of RbfA. However, the structural basis of the mtRbfA-mediated maturation process is poorly understood. Herein, we report the 1H, 15N, and 13C resonance assignments of the KH domain of mtRbfA and its solution structure. The mtRbfA domain adopts essentially the same α1–β1–β2–α2(kinked)–β3 topology as the type-II KH domain. Comparison with the RbfA counterpart showed structural differences in specific regions that function as a putative RNA-binding site. Particularly, the α2 helix of mtRbfA forms a single helix with a moderate kink at the Ser-Ala-Ala sequence, whereas the corresponding α2 helix of RbfA is interrupted by a distinct kink at the Ala-x-Gly sequence, characteristic of bacterial RbfA proteins, to adopt an α2-kink-α3 conformation. Additionally, the region linking α1 and β1 differs considerably in the sequence and structure between RbfA and mtRbfA. These findings suggest some variations of the RNA-binding mode between them and provide a structural basis for mtRbfA function in mitoribosome biogenesis.
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Data availability
The chemical shift assignments for the type-II KH domain of mtRbfA were deposited in the BMRB database with Accession Number 36478. The atomic coordinates for the ensemble of 20 NMR structures calculated by CYANA 2.1 were deposited in the Protein Data Bank (PDB) under the Accession Code 2E7G and those with Amber12 refinement under Accession Code 7X9U, respectively.
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Acknowledgements
We thank Dr. Sumio Sugano in the Department of Medical Genome Sciences, Graduate School of Frontier Sciences, the University of Tokyo, for providing the cDNA clone of mtRbfA, and Mr. M. Furue in the RIKEN Systems and Structural Biology Center for the supporting NMR data analysis.
Funding
This work was supported by the RIKEN Structural Genomics/Proteomics Initiative (RSGI) and by the National Project on Protein Structural and Functional Analyses of the Ministry of Education, Culture, Sports, Science, and Technology of Japan. This work was also supported by grants from the Musashino University Gakuin Tokubetsu Kenkyuhi to YM.
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SW, AT, TT, and MS prepared protein samples. KK, SS, MT, KT, and TN, NK and PG contributed to NMR experiments and structure determination. NN, MT, YM, and KK wrote the paper. YM and SY supervised the study.
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Kuwasako, K., Suzuki, S., Nameki, N. et al. 1H, 13C, and 15N resonance assignments and solution structures of the KH domain of human ribosome binding factor A, mtRbfA, involved in mitochondrial ribosome biogenesis. Biomol NMR Assign 16, 297–303 (2022). https://doi.org/10.1007/s12104-022-10094-3
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DOI: https://doi.org/10.1007/s12104-022-10094-3