tailieunhanh - Báo cáo Y học: Mammalian HSP60 is quickly sorted into the mitochondria under conditions of dehydration

There are few reports concerning the sortingmechanisms of mammalian HSP60 into the mitochondria from the cyto-plasm. In the present study we investigated the protein import system. Based on immunoblotting and immuno-histochemistry, HSP60 was detected in both the cytoplasm andmitochondria. The purified cytoplasmicHSP60 showed chaperone activity, and the protein was imported into the mitochondriain vitro by a mitochondrial import assay. | Eur. J. Biochem. 269 5931-5938 2002 FEBS 2002 doi Mammalian HSP60 is quickly sorted into the mitochondria under conditions of dehydration Hideaki Itoh1 Atsushi Komatsuda2 Hiroshi Ohtani2 Hideki Wakui2 Hirokazu Imai2 Ken-ichi Sawada2 Michiro Otaka3 Masahito Ogura1 Akira Suzuki1 and Fumio Hamada4 1 Department of Biochemistry 2Department of Third Internal Medicine and 3Department of First Internal Medicine Akita University School of Medicine Akita City Japan 4Department of Material-Process Engineering and Applied Chemistry for Environment Akita University Faculty of Engineering and Resource Science Akita City Japan There are few reports concerning the sorting mechanisms of mammalian HSP60 into the mitochondria from the cytoplasm. In the present study we investigated the protein import system. Based on immunoblotting and immunohistochemistry HSP60 was detected in both the cytoplasm and mitochondria. The purified cytoplasmic HSP60 showed chaperone activity and the protein was imported into the mitochondria in vitro by a mitochondrial import assay. HSP60 mRNA was increased in the kidney papilla of rats that had been water restricted for three and five days but no changes in HSP60 mRNA were detected in the cortex or the medulla of the rat kidneys. Upon immunoblotting HSP60 was detected in both the cytoplasm and the mitochondria of normal rat kidney cortex medulla and papilla in almost the same quantity. HSP60 was remarkably decreased in the kidney papilla of rats that were water restricted but the protein was increased in the mitochondria of the rat kidney papilla. We also analysed binding of the protein to the signal sequence of HSP60 using signal sequence-affinity column chromatography. We identified only one protein band with a molecular mass of 70 kDa on SDS PAGE. The protein was eluted from the affinity column by an excess of signal peptide or by 5 mM ATP. Upon immunoblotting the 70-kDa protein cross-reacted with an antibody against .