4Y27

E.coli 23S Sarcin-Ricil Loop, modified with a 2-Me on G2661 and a methylphosphonate on A2662

4Y27 の概要

• エントリーDOI: 10.2210/pdb4y27/pdb
• 関連するPDBエントリー: 3DVZ
• 分子名称: 27-mer 23S Sarcin-Ricil Loop (2 entities in total)
• 機能のキーワード: rna, modification, ribosome
• 由来する生物種: Escherichia coli
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 8756.31

構造登録者

Ennifar, E.,Micura, R.,Fluer, S. (登録日: 2015-02-09, 公開日: 2015-11-18, 最終更新日: 2024-01-10)

主引用文献

Koch, M.,Flur, S.,Kreutz, C.,Ennifar, E.,Micura, R.,Polacek, N.
Role of a ribosomal RNA phosphate oxygen during the EF-G-triggered GTP hydrolysis.
Proc.Natl.Acad.Sci.USA, 112:E2561-E2568, 2015

PubMed Abstract: Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

PubMed: 25941362
DOI: 10.1073/pnas.1505231112

実験手法

X-RAY DIFFRACTION (0.998 Å)