Improved Catenated Structures of Bovine Peroxiredoxin III F190L Reveal Details of Ring-Ring Interactions and a Novel Conformational State.

Zhenbo Cao,Donna P Mcgow,Colin Shepherd,J Gordon Lindsay,Andrew C Gill

doi:10.1371/journal.pone.0123303

Abstract

Mitochondrial 2-cys peroxiredoxin III (PrxIII) is a key player in antioxidant defence reducing locally-generated H2O2 to H2O. A Phe to Leu (F190L) mutation in the C-terminal α-helix of PrxIII, mimicking that found in some bacteria and parasites, increases its resistance to hyperoxidation but has no obvious influence on peroxidase activity. Here we report on the oxidized and reduced crystal structures of bovine PrxIII F190L at 2.4 Å and 2.2 Å, respectively. Both structures exist as two-ring catenanes with their dodecameric rings inclined at 55o to each other, similar to that previously reported for PrxIII C168S. The new higher-resolution structures reveal details of the complex network of H-bonds stabilising the inter-toroid contacts. In addition, Arg123, the key conserved residue, that normally interacts with the catalytic cys (Cp, cys 47) is found in a distinct conformation extending away from the Cp while the characteristic Arg-Glu-Arg network, underpinning the active-site geometry also displays a distinctive arrangement, not observed previously. This novel active-site organisation may provide new insights into the dynamics of the large-scale conformational changes occurring between oxidized and reduced states.

Highlights

Mitochondria are the powerhouses of the cell and the major intracellular sites of reactive oxygen species (ROS) production [1]
ROS are best known for their damaging effects on cellular macromolecules during oxidative stress, there is increasing evidence to indicate that oxidizing agents such as H2O2 play vital roles in redox signalling [2]
We report on the crystal structures of both oxidized and reduced forms of bovine peroxiredoxin III (PrxIII) F190L to 2.4 Å and 2.2 Å, respectively and examine some of its novel structural and enzymatic properties

Summary

Introduction

Mitochondria are the powerhouses of the cell and the major intracellular sites of reactive oxygen species (ROS) production [1]. ROS are best known for their damaging effects on cellular macromolecules during oxidative stress, there is increasing evidence to indicate that oxidizing agents such as H2O2 play vital roles in redox signalling [2]. During respiration linked ATP production in the mitochondrial inner membrane, there is significant electron leakage from the electron transport chain, especially from complexes I and complex III, initially generating superoxide anions (O2.-). Most superoxide is reduced to H2O2 by the mitochondrial Mn2+-requiring superoxide dismutase (MnSOD). Competitive kinetic studies have estimated that 90% of mitochondrial H2O2 is further reduced to water by PLOS ONE | DOI:10.1371/journal.pone.0123303. Competitive kinetic studies have estimated that 90% of mitochondrial H2O2 is further reduced to water by PLOS ONE | DOI:10.1371/journal.pone.0123303 April 23, 2015

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