Kinetic Constraints in the Specific Interaction between Phosphorylated Ubiquitin and Proteasomal Shuttle Factors.

Ling-Yun Qin,Zhou Gong,Kan Liu,Chun Tang,Xu Dong

doi:10.3390/biom11071008

Abstract

Ubiquitin (Ub) specifically interacts with the Ub-associating domain (UBA) in a proteasomal shuttle factor, while the latter is involved in either proteasomal targeting or self-assembly coacervation. PINK1 phosphorylates Ub at S65 and makes Ub alternate between C-terminally relaxed (pUbRL) and retracted conformations (pUbRT). Using NMR spectroscopy, we show that pUbRL but not pUbRT preferentially interacts with the UBA from two proteasomal shuttle factors Ubqln2 and Rad23A. Yet discriminatorily, Ubqln2-UBA binds to pUb more tightly than Rad23A does and selectively enriches pUbRL upon complex formation. Further, we determine the solution structure of the complex between Ubqln2-UBA and pUbRL and uncover the thermodynamic basis for the stronger interaction. NMR kinetics analysis at different timescales further suggests an indued-fit binding mechanism for pUb-UBA interaction. Notably, at a relatively low saturation level, the dissociation rate of the UBA-pUbRL complex is comparable with the exchange rate between pUbRL and pUbRT. Thus, a kinetic constraint would dictate the interaction between Ub and UBA, thus fine-tuning the functional state of the proteasomal shuttle factors.

Highlights

Though the Chemical shift perturbations (CSPs) magnitude is similar for pUbRL when tititrated with two Ub-associating domain (UBA) domains, the largest perturbed residues are found in β4 and β2 trated with two UBA domains, the largest perturbed residues are found in β4 and β2 with with the Ubqln2-UBA β5 and with the Rad23A-UBA2 (Figure 1)
Proteins, the post-transitional modification process known as ubiquitination
An important signaling molecule between and performs its function by modifying namely pUb and pUb titrations, we have shown that the proteins, the post-transitional modification process known as ubiquitination

Summary

Introduction

Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. The ubiquitin-proteasomal system is essential for maintaining proteostasis in the cell. Substrate proteins conjugated with the ubiquitin (Ub) chain in a particular manner can be targeted to the proteasome for degradation. The targeting process is mediated by the interactions between Ub and the intrinsic Ub receptors in the proteasome [1,2,3]

Methods

Results

Conclusion