5THE

Crystal structure of the C-terminal lobe of a budding yeast Argonaute

5THE の概要

• エントリーDOI: 10.2210/pdb5the/pdb
• 分子名称: Uncharacterized protein, RNA (5'-R(P*UP*AP*AP*AP*AP*AP*AP*A)-3') (3 entities in total)
• 機能のキーワード: rna-binding protein, rna binding protein-rna complex, rna binding protein/rna
• 由来する生物種: Vanderwaltozyma polyspora; 詳細
• タンパク質・核酸の鎖数: 8
• 化学式量合計: 254475.28

構造登録者

Dayeh, D.M.,Nakanishi, K. (登録日: 2016-09-29, 公開日: 2018-03-14, 最終更新日: 2023-10-04)

主引用文献

Dayeh, D.M.,Kruithoff, B.C.,Nakanishi, K.
Structural and functional analyses reveal the contributions of the C- and N-lobes of Argonaute protein to selectivity of RNA target cleavage.
J. Biol. Chem., 293:6308-6325, 2018

PubMed Abstract: Some gene transcripts have cellular functions as regulatory noncoding RNAs. For example, ∼23-nucleotide (nt)-long siRNAs are loaded into Argonaute proteins. The resultant ribonucleoprotein assembly, the RNA-induced silencing complex (RISC), cleaves RNAs that are extensively base-paired with the loaded siRNA. To date, base complementarity is recognized as the major determinant of specific target cleavage (or slicing), but little is known about how Argonaute inspects base pairing before cleavage. A hallmark of Argonaute proteins is their bilobal structure, but despite the significance of this structure for curtailing slicing activity against mismatched targets, the molecular mechanism remains elusive. Here, our structural and functional studies of a bilobed yeast Argonaute protein and its isolated catalytic C-terminal lobe (C-lobe) revealed that the C-lobe alone retains almost all properties of bilobed Argonaute: siRNA-duplex loading, passenger cleavage/ejection, and siRNA-dependent RNA cleavage. A 2.1 Å-resolution crystal structure revealed that the catalytic C-lobe mirrors the bilobed Argonaute in terms of guide-RNA recognition and that all requirements for transitioning to the catalytically active conformation reside in the C-lobe. Nevertheless, we found that in the absence of the N-terminal lobe (N-lobe), target RNAs are scanned for complementarity only at positions 5-14 on a 23-nt guide RNA before endonucleolytic cleavage, thereby allowing for some off-target cleavage. Of note, acquisition of an N-lobe expanded the range of the guide RNA strand used for inspecting target complementarity to positions 2-23. These findings offer clues to the evolution of the bilobal structure of catalytically active Argonaute proteins.

PubMed: 29519815
DOI: 10.1074/jbc.RA117.001051

実験手法

X-RAY DIFFRACTION (2.097 Å)