6L2L

The structure of the tRNA-specific deaminase from M. capricolum

6L2L の概要

• エントリーDOI: 10.2210/pdb6l2l/pdb
• 分子名称: Nucleoside deaminase family protein, ZINC ION (3 entities in total)
• 機能のキーワード: trna-specific deaminase, protein engineering, rna binding protein
• 由来する生物種: Mycoplasma capricolum subsp. capricolum
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 18287.46

構造登録者

Xie, W.,Liu, H.,Wu, S. (登録日: 2019-10-05, 公開日: 2020-08-12, 最終更新日: 2023-11-22)

主引用文献

Liu, H.,Wu, S.,Ran, D.,Xie, W.
Structure of a tRNA-specific deaminase with compromised deamination activity.
Biochem.J., 477:1483-1497, 2020

PubMed Abstract: Nucleotide 34 in tRNA is extensively modified to ensure translational fidelity and efficacy in cells. The deamination of adenosine at this site catalyzed by the enzyme TadA gives rise to inosine (I), which serves as a typical example of the wobble hypothesis due to its diverse basepairing capability. However, recent studies have shown that tRNAArgACG in Mycoplasma capricolum contains unmodified adenosine, in order to decode the CGG codon. The structural basis behind the poorly performing enzyme M. capricolum TadA (McTadA) is largely unclear. Here we present the structures of the WT and a mutant form of McTadA determined at high resolutions. Through structural comparison between McTadA and other active TadA enzymes as well as modeling efforts, we found that McTadA presents multiple structural conflicts with RNA substrates and thus offered support to previous studies from a structural perspective. These clashes would potentially lead to reduced substrate binding affinity of McTadA, consistent with our in vitro deamination activity and binding assays. To rescue the deamination activity of McTadA, we carried out two rounds of protein engineering through structure-guided design. The unsuccessful attempts of the activity restoration could be attributed to the altered dimer interface and stereo hindrance from the non-catalytic subunit of McTadA, which could be the inevitable outcome of the natural evolution. Our study provides structural insight into an alternative decoding and evolutionary strategy by a compromised TadA enzyme at a molecular level.

PubMed: 32270856
DOI: 10.1042/BCJ20190858

実験手法

X-RAY DIFFRACTION (2.40045827641 Å)