Monitoring and analysis of mitochondrial precursor protein aggregates in the cytosol.

The vast majority of mitochondrial precursor proteins is synthesized in the cytosol and subsequently imported into the organelle with the help of targeting signals that are present within these proteins. Disruptions in mitochondrial import will result in the accumulation of the organellar precursors in the cytosol of the cell. If mislocalized proteins exceed their critical concentrations, they become prone to aggregation.

Methods to monitor mitochondrial disulfide bonds.

Mitochondria contain numerous proteins that utilize the chemistry of cysteine residues, which can be reversibly oxidized. These proteins are involved in mitochondrial biogenesis, protection against oxidative stress, metabolism, energy transduction to adenosine triphosphate, signaling and cell death among other functions. Many proteins located in the mitochondrial intermembrane space are imported by the mitochondrial import and assembly pathway the activity of which is based on the reversible oxidation of cysteine residues and oxidative trapping of substrates.

Temporal alterations of the nascent proteome in response to mitochondrial stress.

Under stress, protein synthesis is attenuated to preserve energy and mitigate challenges to protein homeostasis. Here, we describe, with high temporal resolution, the dynamic landscape of changes in the abundance of proteins synthesized upon stress from transient mitochondrial inner membrane depolarization. This nascent proteome was altered when global translation was attenuated by stress and began to normalize as translation was recovering.

Structure prediction analysis of human core TIM23 complex reveals conservation of the protein translocation mechanism.

The majority of mitochondrial proteins are encoded in the nucleus, translated on cytosolic ribosomes, and subsequently targeted to the mitochondrial surface. Their further import into the organelle is facilitated by highly specialized protein translocases. Mitochondrial precursor proteins that are destined to the mitochondrial matrix and, to some extent, the inner membrane, utilize translocase of the inner membrane (TIM23).