Redox regulation of protein folding in the mitochondrial intermembrane space

Abstract

Protein translocation pathways to the mitochondrial matrix and inner membrane have been well characterized. However, translocation into the intermembrane space, which was thought to be simply a modification of the traditional translocation pathways, is complex. The mechanism by which a subset of intermembrane space proteins, those with disulfide bonds, are translocated has been largely unknown until recently. Specifically, the intermembrane space proteins with disulfide bonds are imported via the mitochondrial intermembrane space assembly (MIA) pathway. Substrates are imported via a disulfide exchange relay with two components Mia40 and Erv1. This new breakthrough has resulted in novel concepts for assembly of proteins in the intermembrane space, suggesting that this compartment may be similar to that of the endoplasmic reticulum and the prokaryotic periplasm. As a better understanding of this pathway emerges, new paradigms for thiol-disulfide exchange mechanisms may be developed. Given that the intermembrane space is important for disease processes including apoptosis and neurodegeneration, new roles in regulation by oxidation-reduction chemistry seem likely to be relevant.

Fig. 1

Schematic of translocation and assembly complexes in the mitochondrion. Nuclear-coded mitochondrial precursors cross the TOM complex. The SAM complex mediates the assembly of outer membrane proteins with complicated topologies. Proteins with a typical N-terminal targeting presequence are imported via the TIM23 complex, whereas inner membrane proteins such as the carrier proteins are imported via the TIM22 complex.

Fig. 2

The Mia40-Erv1 import pathway for CX₃C and CX₉C proteins. The substrates (model substrate is a small Tim protein) are imported in an unfolded, reduced state across the outer membrane (OM). Oxidized Mia40 serves as the import receptor and forms a transient disulfide with the substrate. The substrate leaves in an oxidized state and a series of disulfide exchange reactions continue through Erv1 C130–C133 to FAD. From FADH₂, the electrons are passed to different acceptors. (A) Electrons may be shuttled to cytochrome c and then to the electron transport chain (ETS) or oxidized Ccp1. (B) Alternatively, electrons may be shuttled to oxygen to generate hydrogen peroxide, which is removed by reduced Ccp1. Ccp1 and cyt c can function together to bypass the requirement for the electron transport system. The number of electrons (e⁻) that are transferred at each step are indicated. Details have not been elucidated for the specific exchange reactions and additional components most likely participate in this pathway.

Fig. 3

The import pathway for carrier proteins is redox regulated. In the assembled Tim9–Tim10 complex, the small Tim proteins are oxidized and bind to substrate as it emerges from the TOM complex. In the presence of oxidant, a translocation intermediate bound to the small Tim complexes can be arrested in the intermembrane space (Stage 1). In the presence of reductant, the translocation intermediate is subsequently “chased” and the substrate inserts into the inner membrane (Stage 2). The small Tim proteins are reassembled for another round of import, which may be facilitated by Hot1p (HOT; Stage 3). Detailed mechanistic steps have not been elucidated and additional proteins may participate in this pathway.