The mitochondrial intermembrane space (IMS) contains many small cysteine-bearing proteins, and their passage across the outer membrane and subsequent folding require recognition and disulfide bond transfer by an oxidative translocator Tim40/Mia40 in the inner membrane facing the IMS. Here we determined the crystal structure of the core domain of yeast Mia40 (Mia40C4) as a fusion protein with maltose-binding protein at a resolution of 3 Å. The overall structure of Mia40C4 is a fruit-dish-like shape with a hydrophobic concave region, which accommodates a linker segment of the fusion protein in a helical conformation, likely mimicking a bound substrate. Replacement of the hydrophobic residues in this region resulted in growth defects and impaired assembly of a substrate protein. The Cys296-Cys298 disulfide bond is close to the hydrophobic concave region or possible substrate-binding site, so that it can mediate disulfide bond transfer to substrate proteins. These results are consistent with the growth phenotypes of Mia40 mutant cells containing Ser replacement of the conserved cysteine residues.

Mitochondria import most of their resident proteins from the cytosol, and the import receptor Tom20 of the outer-membrane translocator TOM40 complex plays an essential role in specificity of mitochondrial protein import. Here we analyzed the effects of Tom20 binding on NMR spectra of a long mitochondrial presequence and found that it contains two distinct Tom20-binding elements. In vitro import and cross-linking experiments revealed that, although the N-terminal Tom20-binding element is essential for targeting to mitochondria, the C-terminal element increases efficiency of protein import in the step prior to translocation across the inner membrane. Therefore Tom20 has a dual role in protein import into mitochondria: recognition of the targeting signal in the presequence and tethering the presequence to the TOM40 complex to increase import efficiency.protein's function relies on its correct subcellular location. Newly synthesized proteins are delivered to their sites of actions by cellular protein transport systems. Because subcellular compartments are bounded by biological membrane(s) in eukaryotic cells, the protein transport needs to start with insertion into or translocation across the target membrane. Therefore most protein transport systems involve a targeting signal on the cargo protein and a multiprotein complex on the target membrane called a translocator (1).Mitochondria are essential organelles in eukaryotic cells that consist of the outer and inner membranes and two aqueous compartments, the intermembrane space (IMS) and the matrix. Most mitochondrial proteins are encoded in the nuclear genome, synthesized in the cytosol, and subsequently imported into mitochondria. Mitochondrial protein import is mediated by translocators in the outer and inner membranes, including two TOM (translocase of the outer mitochondrial membrane) complexes, the TOM40 complex and the TOB (topogenesis of outer-membrane β-barrel proteins)/SAM (sorting and assembly machinery) complex, and two TIM (translocase of the inner mitochondrial membrane) complexes, the TIM23 complex and the TIM22 complex (2-5). The import pathway generally starts from the TOM40 complex and then branches out into several distinct intramitochondrial sorting pathways involving other translocators.The targeting information for mitochondria is contained in the N-terminal cleavable presequence or within the mature part of precursor proteins. The targeting information as well as intramitochondrial sorting information is recognized by several receptor subunits of the TOM and TIM complexes along the import pathways. Among them, Tom20, a peripheral subunit of the TOM40 complex, is a general import receptor that recognizes mitochondrial targeting signals contained in presequences. Tom20 is anchored to the outer membrane by the N-terminal hydrophobic segment and exposes a receptor domain to the cytosol. The NMR and X-ray structures of the receptor domain of rat Tom20 in a complex with a presequence peptide showed that the bound presequence forms an amphiphilic helix...

Here, we report the identification of yeast 15-kD Tim15/Zim17, a new member of mitochondrial Hsp70 (mtHsp70)-associated motor and chaperone (MMC) proteins. The 15-kD MMC protein is a peripheral inner membrane protein with a zinc-finger motif. Depletion of the 15-kD protein led to impaired import of presequence-containing proteins into the matrix in vivo and in vitro. Overexpression of the 15-kD protein rescued the functional defects of mtHsp70 in ssc1-3 cells, and a fusion protein containing the 15-kD protein physically interacts with purified mtHsp70. Tim15/Zim17 therefore cooperates with mtHsp70 to facilitate import of presequence-containing proteins into the matrix.

Mitochondrial heat-shock protein 70 (mtHsp70) and its partner proteins drive protein import into the matrix. Tim15/Zim17/ Hep1 is a mtHsp70 partner protein on the matrix side of the inner mitochondrial membrane. We determined the nuclear magnetic resonance (NMR) structure of the core domain of Tim15. On the basis of the NMR structure, we created Tim15 mutants and tested their ability to complement the functional defects of Tim15 depletion and to suppress self-aggregation of mtHsp70 in vivo. A pair of basic residues, Arg 106 and His 107, conserved Asp 111 and flexible loop 133-137, and were important (Arg 106-His 107 pair and Asp 111) or partly important (the loop 133-137) for yeast cell growth, mitochondrial protein import and the suppression of mtHsp70 aggregation. Therefore, the function of Tim15 in yeast cell growth is well correlated with its ability to suppress mtHsp70 aggregation, although it is still unknown whether inhibition of mtHsp70 aggregation is the primary function of Tim15.

Enormous quantities of radionuclides were released into the environment following the disastrous accident at the Fukushima Daiichi Nuclear Power Plant (FDNPP) in March 2011. It is of great importance to determine the exposure doses received by the populations living in the radiologically affected areas; however, there has been significant difficulty in estimating the internal thyroid dose received through the intake of short-lived radionuclides (mainly, 131I), because of the lack of early measurements on people. An estimation by the National Institute of Radiological Sciences for 1 April 2012 to 31 March 2013 was thus performed using a combination of the following three sources: thyroid measurement data (131I) for 1080 children examined in the screening campaign, whole-body counter measurement data (134Cs, 137Cs) for 3000 adults, and atmospheric transport dispersion model simulations. In this study, the residents of Futaba town, Iitate village and Iwaki city were shown to have the highest thyroid equivalent dose, and their doses were estimated to be mostly below 30 mSv. However, this result involved a lot of uncertainties and provided only representative values for the residents. The present paper outlines a more recent dose estimation and preliminary analyses of personal behavior data used in the new method.