Journal of Biological Chemistry
Volume 287, Issue 42, 12 October 2012, Pages 34961-34969
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Protein Structure and Folding
Structure of Yeast Sulfhydryl Oxidase Erv1 Reveals Electron Transfer of the Disulfide Relay System in the Mitochondrial Intermembrane Space*

https://doi.org/10.1074/jbc.M112.394759Get rights and content
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The disulfide relay system in the mitochondrial intermembrane space drives the import of proteins with twin CX9C or twin CX3C motifs by an oxidative folding mechanism. This process requires disulfide bond transfer from oxidized Mia40 to a substrate protein. Reduced Mia40 is reoxidized/regenerated by the FAD-linked sulfhydryl oxidase Erv1 (EC 1.8.3.2). Full-length Erv1 consists of a flexible N-terminal shuttle domain (NTD) and a conserved C-terminal core domain (CTD). Here, we present crystal structures at 2.0 Å resolution of the CTD and at 3.0 Å resolution of a C30S/C133S double mutant of full-length Erv1 (Erv1FL). Similar to previous homologous structures, the CTD exists as a homodimer, with each subunit consisting of a conserved four-helix bundle that accommodates the isoalloxazine ring of FAD and an additional single-turn helix. The structure of Erv1FL enabled us to identify, for the first time, the three-dimensional structure of the Erv1NTD, which is an amphipathic helix flanked by two flexible loops. This structure also represents an intermediate state of electron transfer from the NTD to the CTD of another subunit. Comparative structural analysis revealed that the four-helix bundle of the CTD forms a wide platform for the electron donor NTD. Moreover, computational simulation combined with multiple-sequence alignment suggested that the amphipathic helix close to the shuttle redox enter is critical for the recognition of Mia40, the upstream electron donor. These findings provide structural insights into electron transfer from Mia40 via the shuttle domain of one subunit of Erv1 to the CTD of another Erv1 subunit.

Cysteine-mediated Cross-linking
Electron Transfer
Oxidation-Reduction
Sulfhydryl
X-ray Crystallography
Yeast
Disulfide Relay System

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The atomic coordinates and structure factors (codes 4E0H and 4E0I) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

*

This work was supported by Ministry of Science and Technology of China Project 2012CB911000.