Zinc Can Play Chaperone-like and Inhibitor Roles during Import of Mitochondrial Small Tim Proteins

Bruce Morgan,Swee Kim Ang,Guanhua Yan,Hui Lu

doi:10.1074/jbc.m808691200

Abstract

Zinc is an essential cofactor required for the function of approximately 8% of the yeast and 10% of the human proteome. All of the "small Tim" proteins of the mitochondrial intermembrane space contain a strictly conserved "twin CX(3)C" zinc finger motif, which can bind zinc ions in the Cys-reduced form. We have shown previously that although disulfide bond formation is essential for the function of these proteins in mitochondria, only reduced proteins can be imported into mitochondria (Lu, H., Allen, S., Wardleworth, L., Savory, P., and Tokatlidis, K. (2004) J. Biol. Chem. 279, 18952-18958 and Morgan, B., and Lu, H. (2008) Biochem. J. 411, 115-122). However, the role of zinc during the import of these proteins is unclear. This study shows that the function of zinc is complex. It can play a thiol stabilizer role preventing oxidative folding of the small Tim proteins and maintaining the proteins in an import-competent form. On the other hand, zinc-bound forms cannot be imported into mitochondria efficiently. Furthermore, our results show that zinc is a powerful inhibitor of Erv1, an essential component of the import pathway used by the small Tim proteins. We propose that zinc plays a chaperone-like role in the cytosol during biogenesis of the small Tim proteins and that the proteins are imported into mitochondria through the apo-forms.

Highlights

The small Tim proteins of the Saccharomyces cerevisiae yeast mitochondrial intermembrane space (IMS)2 play an essential role during the import of mitochondrial membrane proteins [14, 15], and they are themselves imported through the redoxregulated Mia40/Erv1 pathway [16, 17]
Using mitochondrial import coupled with a buffered zinc system, our results show that whereas the precursor Tim9 was stabilized in the reduced form, the level of mitochondrial import was decreased with the increase of free zinc concentration
The reduced small Tim apoproteins can be oxidized under the cytosolic redox conditions, which inhibit their import into mitochondria

Summary

Introduction

The small Tim proteins of the Saccharomyces cerevisiae yeast mitochondrial intermembrane space (IMS)2 play an essential role during the import of mitochondrial membrane proteins [14, 15], and they are themselves imported through the redoxregulated Mia40/Erv1 pathway [16, 17]. To understand whether ZnTim9 can be imported efficiently, was oxidized by GSSG under cytosolic redox conditions with conditions at which most (if not all) proteins can be stably oxidized Tim9 observed after ϳ5 min, and the oxidative folding maintained in the zinc-bound form are required.

Results

Conclusion