Structural Analysis of the TIM23 Complex Sheds light on the Intricate Protein-Protein Interactions of the Mitochondrial Import System

Mitochondria are essential organelles hosting several vital cellular functions, such as: providing cellular energy, Iron-Sulfur cluster assembly and controlling programmed cell death. These functions are dictated by ~1000 mitochondrial proteins, most of which are synthesized in the cytoplasm and must be translocated to their correct sub-mitochondrial compartments. This process is mediated by several intricate mitochondrial protein-targeting systems.

Our research goal was to determine the function mechanism of one of the major mitochondrial protein import complexes - TIM23, using structural analysis methods. To achieve this goal, we chose two different strategies: Initially, attempts to solve the crystal structure of parts of the complex, which failed. Next, we designed a laser-printed, peptide macro-array chip, containing thousands of dissected sequence segments of TIM23 components, including mutation combinations of important amino acids. In a series of experiments, we analyzed the interaction patterns of purified TIM23 components to the array of peptides, and managed to fully characterize several important interactions with an innovative, unique and very quick technique. Using the information gathered from the peptide-array, we managed to predict an in-silico model of some of the complex components. Thus, providing vital insights on the intricate inter-complex cooperation required for proper translocation of proteins into the mitochondrion.

Recently, mutations in Tim50, one of TIM23 components, were associated with a severe neurological disorder, found in children from inbred families. In the future, we plan to utilize our gathered knowledge to try and identify if the disease phenotypes are related to Tim50 function in the mitochondrial import process.