Protein Export across the Inner Membrane of Mitochondria

Abstract

The biogenesis of mitochondria requires the insertion of both nuclear and mitochondrially encoded proteins into the inner membrane. The inner membrane protein Oxa1 plays an important role in this process. Translocation of the terminal intermembrane space domains of subunit 2 of the cytochrome oxidase complex, Cox2, strictly depends on Oxa1. In contrast, other Oxa1 substrates can be inserted independently of Oxa1 function, although at reduced efficiency. A Saccharomyces cerevisiae mutant containing a large deletion in its mitochondrial genome allowed us to analyze the insertion process of a fusion protein of cytochrome b and Cox2. In this mutant, the N-terminal domain of Cox2 is synthesized as a hairpin loop that is flanked by hydrophobic transmembrane segments on both sides. Both genetic and biochemical evidences indicate that translocation of this region across the inner membrane still requires Oxa1 function. Thus, the position of intermembrane space domains within protein sequences does not appear to determine their dependence on the Oxa1 translocase. Our observations rather suggest that the dependence on Oxa1 correlates with the net charge of the domain that has to be translocated across the lipid bilayer.

Determinants for Stop-transfer and Post-import Pathways for Protein Targeting to the Chloroplast Inner Envelope Membrane

“…In mitochondria, similar stop-transfer and post-import pathways for inner membrane insertion exist (33,34), and it has been shown that the presence of prolines in the TMD is a fundamental determinant in the capability of a TMD to be arrested or not in the membrane during translocation. It has been demonstrated that prolines in the TMD cause these helices to be transferred by the translocon to the matrix, disfavoring TMD arrest and transfer to the lipid bilayer (35) and prompting these proteins to be inserted via a conservative sorting process that is similar to the post-import pathway in chloroplasts.…”

Mechanisms of YidC-mediated Insertion and Assembly of Multimeric Membrane Protein Complexes

“…This suggests that the negative charges in the loop region are not essential for YidC-mediated translocation and that topogenesis in the YidC-only pathway is dictated mostly by the positively charged residues remaining in the cytosol. In contrast, the charges present in the translocated regions of substrates determine the Oxa1p dependence (37).…”

A Role for Pet100p in the Assembly of Yeast Cytochrome c Oxidase

“…Although the mitochondrial import pathways have been elucidated (for review, see Ref. 15) and the mitochondrial export pathway is beginning to yield to analysis (16), the assembly pathway for cytochrome c oxidase is still poorly understood. However, it is now clear that assembly of cytochrome c oxidase requires proteins encoded by a number of different nuclear genes (1,(17)(18)(19)(20)(21)(22).…”

“…Among the genes with specific effects on cytochrome c oxidase assembly are three genes (COX10, COX15, and YAH1) required for the synthesis of heme A (25)(26)(27)(28), four genes (SCO1, COX11, COX17, and COX23) required for the recruitment and uptake of copper into mitochondria and its assembly into holocytochrome c oxidase (29 -37), and one gene (OXA1/PET1402) that is part of a translocation channel involved in mitochondrial export (16). In addition to the above genes, there are at least five other genes (PET100, PET117, PET191, COX14, and COX16) (38 -42) that encode proteins that have been proposed to function as "assembly facilitators" (1).…”