PMID: 9419343Feb 21, 1998Paper

Characterization of mammalian translocase of inner mitochondrial membrane (Tim44) isolated from diabetic newborn mouse kidney

Proceedings of the National Academy of Sciences of the United States of America
J Wada, Yashpal S Kanwar

Abstract

Mammalian translocase of mitochondrial inner membrane (mTim44) was isolated during representational difference analysis of cDNA from diabetic mouse kidney. Streptozotocin-induced diabetic mouse kidney cDNA was prepared and subtracted by normal mouse kidney cDNA. By using one of the isolated cDNA fragments as a screening probe, full-length cDNA of mTim44 was isolated from lambdaZAP kidney cDNA library. At the nucleotide level, mTim44 did not exhibit significant homology with any known genes; however, at the amino acid level, it had 50% similarity and 29% identity with yeast Tim44. C-terminal FLAG epitope-tagged mTim44 fusion protein was transiently expressed in COS7 cells. By using anti-FLAG epitope M2 monoclonal antibody, mTim44 was found to have its subcellular localization associated with mitochondria. By immunoelectron microscopy, mTim44 was seen in the paracrystalline structures within the mitochondria, as well as in their cristae. Mitochondrial import assay of in vitro translated mTim44 indicated that its precursor product ( approximately 50 kDa) was imported and proteolytically processed to a mature approximately 44-kDa protein, and its translocation was inner membrane potential (DeltaPsi)-dependent. Imported mTim44 was p...Continue Reading

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