Structural and functional characterization of the mitochondrial complex IV assembly factor Coa6.

Shadi Maghool,N Dinesha G Cooray,Michael T Ryan,David Aragão,Megan J Maher,David A Stroud

doi:10.26508/lsa.201900458

Abstract

Assembly factors play key roles in the biogenesis of many multi-subunit protein complexes regulating their stability, activity, and the incorporation of essential cofactors. The human assembly factor Coa6 participates in the biogenesis of the CuA site in complex IV (cytochrome c oxidase, COX). Patients with mutations in Coa6 suffer from mitochondrial disease due to complex IV deficiency. Here, we present the crystal structures of human Coa6 and the pathogenic W59CCoa6-mutant protein. These structures show that Coa6 has a 3-helical bundle structure, with the first 2 helices tethered by disulfide bonds, one of which likely provides the copper-binding site. Disulfide-mediated oligomerization of the W59CCoa6 protein provides a structural explanation for the loss-of-function mutation.

Highlights

The mitochondrial oxidative phosphorylation (OXPHOS) system generates the bulk of cellular ATP, fuelling the energy demands of most eukaryotes
Recombinant wild-type Coa6 (WTCoa6) protein was overexpressed in Escherichia coli strain SHuffle T7, which promotes the production of correctly disulfide bonded active proteins within the cytoplasm [23, 24]
The COX assembly factor Coa6 was identified through a proteomic survey of the mitochondrial intermembrane space (IMS) from Saccharomyces cerevisiae [49] and the significant number of investigations that followed, proposed its role in the biogenesis of the CuA site, presumably through the interaction of Coa6 with other critical proteins in the COX biogenesis pathway such as Sco1 and Sco2 [10, 11, 21, 51]

Summary

Introduction

The mitochondrial oxidative phosphorylation (OXPHOS) system generates the bulk of cellular ATP, fuelling the energy demands of most eukaryotes. Five multi-subunit protein complexes in the mitochondrial inner membrane, termed complexes I–V, comprise the OXPHOS system. Complex IV (cytochrome c oxidase; COX) is the last complex of the electron transport chain, transferring electrons from cytochrome c to molecular oxygen, and in the process, pumping four protons across the inner membrane [1]. Cytochrome c docks onto the intermembrane space (IMS) domain of COX2, which contains a binuclear copper center, termed CuA that accepts the electrons. The electrons are passed to a heme a group in COX1 and to a heme a3-CuB site and to oxygen, which is reduced to water [5]. Reduction of the CuA site in COX2 represents the critical first stage in complex IV activity

Methods

Results

Conclusion