Chaperone activation and client binding of a 2-cysteine peroxiredoxin

Filipa Teixeira,Karl A T Makepeace,Leslie B Poole,Eric Tse,Ursula Jakob,James C Bardwell,Christoph H Borchers,Ana M Tomás,Daniel R Southworth,Ben A Meinen,Helena Castro

doi:10.1038/s41467-019-08565-8

Abstract

Many 2-Cys-peroxiredoxins (2-Cys-Prxs) are dual-function proteins, either acting as peroxidases under non-stress conditions or as chaperones during stress. The mechanism by which 2-Cys-Prxs switch functions remains to be defined. Our work focuses on Leishmania infantum mitochondrial 2-Cys-Prx, whose reduced, decameric subpopulation adopts chaperone function during heat shock, an activity that facilitates the transition from insects to warm-blooded host environments. Here, we have solved the cryo-EM structure of mTXNPx in complex with a thermally unfolded client protein, and revealed that the flexible N-termini of mTXNPx form a well-resolved central belt that contacts and encapsulates the unstructured client protein in the center of the decamer ring. In vivo and in vitro cross-linking studies provide further support for these interactions, and demonstrate that mTXNPx decamers undergo temperature-dependent structural rearrangements specifically at the dimer-dimer interfaces. These structural changes appear crucial for exposing chaperone-client binding sites that are buried in the peroxidase-active protein.

Highlights

Many 2-Cys-peroxiredoxins (2-Cys-Prxs) are dual-function proteins, either acting as peroxidases under non-stress conditions or as chaperones during stress
The catalytic activity of 2-Cys-peroxiredoxins, which comprise the Prx[1] family[4], is mediated by the active site peroxidatic cysteine Cp, which reacts with peroxide and related oxidants, and undergoes reversible sulfenic acid formation[5]
We recently reported that the mitochondrial 2-Cys Prx of Leishmania infantum adopts two functions, as a peroxidase and as a molecular chaperone[14,15]

Summary

Introduction

Many 2-Cys-peroxiredoxins (2-Cys-Prxs) are dual-function proteins, either acting as peroxidases under non-stress conditions or as chaperones during stress. In vivo and in vitro cross-linking studies provide further support for these interactions, and demonstrate that mTXNPx decamers undergo temperature-dependent structural rearrangements at the dimer-dimer interfaces. These structural changes appear crucial for exposing chaperone-client binding sites that are buried in the peroxidase-active protein. The active site cysteine-containing Cp-loop-helix adopts a closed conformation through an elaborate network of electrostatic interactions, thereby exposing critical aromatic amino acids that pack tightly against the other dimer, stabilizing the dimer–dimer interface (i.e., A-type or alternate interface)[5]. Overoxidation of the active site cysteine to sulfinic acid has been shown to cause the formation of even higher molecular weight oligomeric structures, including filamentous or spherical structures[8,9,10]

Methods

Results

Conclusion