Characterization of the Trimeric TOM Complex by HS-AFM Single-Molecule Analysis.
10 affiliations
- 1Department of Clinical Laboratory Science, Graduate School of Medical Science, Kanazawa University, Kanazawa, Japan.
- 2Department of Biological Sciences, Graduate School of Science, the University of Tokyo, Tokyo, Japan.
- 3Cellular and Molecular Biotechnology Reserach Institute (CMB), National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan.
- 4Integrated Reserach Center for Self-Care Technology (IRC-SRT), National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan.
- 5Department of Biochemistry, Graduate School of Medicine, University of the Ryukyus, Ginowan, Japan.
- 6WPI-Nano Life Science Institute, Kanazawa University, Kanazawa, Japan.
- 7Institute of Science Tokyo, Institute of Integrated Research, Cell Biology Center, Yokohama, Japan.
- 8Faculty of Life Sciences, Kyoto Sangyo University, Kyoto, Japan.
- 9Institute for Protein Dynamics, Kyoto Sangyo University, Kyoto, Japan.
- 10Environmental Stress Research Center (eSRC), Kanazawa University, Ginowan, Japan.
Abstract
The translocase of the outer mitochondrial membrane (TOM) complex is the entry gate for mitochondrial proteins synthesized in the cytosol. Previous biochemical studies indicated that the TOM complex functions as a trimer and exists in a dynamic equilibrium with a Tom22-lacking dimer in intact mitochondria. In contrast, recent cryo-electron microscopy (cryo-EM) studies revealed near-atomic structures of the Tom22-containing core dimer. However, the relationship between the core dimer and the functional trimer remains unclear. Here, we analyzed the oligomeric organization and structural properties of purified yeast TOM complexes using high-speed atomic force microscopy (HS-AFM). Purified yeast TOM complexes predominantly adopted a trimeric organization but underwent deformation and dissociation during repeated HS-AFM scanning, leaving dimeric and monomeric particles. Trimeric particles exhibited spherical shapes consistent with previous crosslinking models, whereas dissociated dimers closely resembled TOM core-dimer structures determined by cryo-EM. HS-AFM also revealed the peripheral localization of the Tom20 receptor and enabled direct visualization of substrate-TOM interactions, with TOM association accompanied by reduced substrate mobility. Together, these findings provide quantitative and structural evidence for a predominantly trimeric organization of the purified TOM complex and suggest that the trimer contains a stable core-dimer unit with a more loosely associated third Tom40-containing unit.
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BibTeX
@article{kobayashi2026characterization,
title = {{Characterization of the Trimeric TOM Complex by HS-AFM Single-Molecule Analysis.}},
author = {Nanako Kobayashi and Satoshi N Omura and Kana Kuzasa and Kenichiro Imai and Shiho Kawai and Hirotatsu Imai and Romain Amyot and Kenichi Umeda and Osamu Nureki and Toshiya Endo and Noriyuki Kodera and Yuhei Araiso},
journal = {Genes to cells : devoted to molecular \& cellular mechanisms},
year = {2026},
month = oct,
doi = {10.1111/gtc.70154},
url = {https://europepmc.org/article/MED/42920373},
note = {CC BY},
}
RIS
TY - JOUR
TI - Characterization of the Trimeric TOM Complex by HS-AFM Single-Molecule Analysis.
AU - Nanako Kobayashi
AU - Satoshi N Omura
AU - Kana Kuzasa
AU - Kenichiro Imai
AU - Shiho Kawai
AU - Hirotatsu Imai
AU - Romain Amyot
AU - Kenichi Umeda
AU - Osamu Nureki
AU - Toshiya Endo
AU - Noriyuki Kodera
AU - Yuhei Araiso
PY - 2026
DA - 2026/10/30
JO - Genes to cells : devoted to molecular & cellular mechanisms
DO - 10.1111/gtc.70154
UR - https://europepmc.org/article/MED/42920373
AB - The translocase of the outer mitochondrial membrane (TOM) complex is the entry gate for mitochondrial proteins synthesized in the cytosol. Previous biochemical studies indicated that the TOM complex functions as a trimer and exists in a dynamic equilibrium with a Tom22-lacking dimer in intact mitochondria. In contrast, recent cryo-electron microscopy (cryo-EM) studies revealed near-atomic structures of the Tom22-containing core dimer. However, the relationship between the core dimer and the functional trimer remains unclear. Here, we analyzed the oligomeric organization and structural properties of purified yeast TOM complexes using high-speed atomic force microscopy (HS-AFM). Purified yeast TOM complexes predominantly adopted a trimeric organization but underwent deformation and dissociation during repeated HS-AFM scanning, leaving dimeric and monomeric particles. Trimeric particles exhibited spherical shapes consistent with previous crosslinking models, whereas dissociated dimers closely resembled TOM core-dimer structures determined by cryo-EM. HS-AFM also revealed the peripheral localization of the Tom20 receptor and enabled direct visualization of substrate-TOM interactions, with TOM association accompanied by reduced substrate mobility. Together, these findings provide quantitative and structural evidence for a predominantly trimeric organization of the purified TOM complex and suggest that the trimer contains a stable core-dimer unit with a more loosely associated third Tom40-containing unit.
SN - 1356-9597
SN - 1365-2443
ER -