Lys63-linked ubiquitin chain adopts multiple conformational states for specific target recognition.

Zhu Liu,Zhou Gong,Wen-Xue Jiang,Wen-Kai Zhu,Wei-Ping Zhang,Chun Tang,Ju Yang,Mai-Li Liu,Da-Chuan Guo

doi:10.7554/elife.05767

Abstract

A polyubiquitin comprises multiple covalently linked ubiquitins and recognizes myriad targets. Free or bound to ligands, polyubiquitins are found in different arrangements of ubiquitin subunits. To understand the structural basis for polyubiquitin quaternary plasticity and to explore the target recognition mechanism, we characterize the conformational space of Lys63-linked diubiquitin (K63-Ub2). Refining against inter-subunit paramagnetic NMR data, we show that free K63-Ub2 exists as a dynamic ensemble comprising multiple closed and open quaternary states. The quaternary dynamics enables K63-Ub2 to be specifically recognized in a variety of signaling pathways. When binding to a target protein, one of the preexisting quaternary states is selected and stabilized. A point mutation that shifts the equilibrium between the different states modulates the binding affinities towards K63-Ub2 ligands. This conformational selection mechanism at the quaternary level may be used by polyubiquitins of different lengths and linkages for target recognition.

Highlights

Ubiquitin is a 76-residue signaling protein found ubiquitously in cells
Our data are consistent with the previous nuclear magnetic resonance (NMR) studies of K63-Ub2 (Tenno et al, 2004; Varadan et al, 2004); in the latter work, the authors failed to detect cross-saturations between the two subunits
We have shown that about 70% of K63-Ub2 exists in the closed state, whereas only about 30% of the protein exists in the open state

Summary

Introduction

Ubiquitin is a 76-residue signaling protein found ubiquitously in cells. Multiple ubiquitins are covalently linked to form a polyubiquitin, which can be attached to a substrate protein. The process is known as ubiquitination, a post-translational modification of the substrate protein. Three classes of enzymes catalyze ubiquitination: ubiquitin-activation enzyme (E1), ubiquitin-conjugation enzymes (E2), and ubiquitin-protein ligases (E3). E2 and E3 dictate ubiquitin linkage and substrate specificities. Deubiquitinases (DUBs) are responsible for erasing ubiquitin signals from a substrate protein (Clague et al, 2012)

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Conclusion