Adaptation of a genetic screen reveals an inhibitor for mitochondrial protein import component Tim44

Non Miyata, Zhiye Tang, Michael A. Conti, Meghan E. Johnson, Colin J. Douglas, Samuel A. Hasson, Robert Damoiseaux, Chia En A. Chang, Carla M. Koehler

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Diverse protein import pathways into mitochondria use translocons on the outer membrane (TOM) and inner membrane (TIM). We adapted a genetic screen, based on Ura3 mistargeting from mitochondria to the cytosol, to identify small molecules that attenuated protein import. Small molecule mitochondrial import blockers of the Carla Koehler laboratory (MB)-10 inhibited import of substrates that require the TIM23 translocon. Mutational analysis coupled with molecular docking and molecular dynamics modeling revealed that MB-10 binds to a specific pocket in the C-terminal domain of Tim44 of the protein-associated motor (PAM) complex. This region was proposed to anchor Tim44 to the membrane, but biochemical studies with MB-10 show that this region is required for binding to the translocating precursor and binding to mtHsp70 in low ATP conditions. This study also supports a direct role for the PAM complex in the import of substrates that are laterally sorted to the inner membrane, as well as the mitochondrial matrix. Thus, MB-10 is the first small molecule modulator to attenuate PAM complex activity, likely through binding to the C-terminal region of Tim44.

Original languageEnglish
Pages (from-to)5429-5442
Number of pages14
JournalJournal of Biological Chemistry
Volume292
Issue number13
DOIs
Publication statusPublished - Mar 31 2017

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Miyata, N., Tang, Z., Conti, M. A., Johnson, M. E., Douglas, C. J., Hasson, S. A., Damoiseaux, R., Chang, C. E. A., & Koehler, C. M. (2017). Adaptation of a genetic screen reveals an inhibitor for mitochondrial protein import component Tim44. Journal of Biological Chemistry, 292(13), 5429-5442. https://doi.org/10.1074/jbc.M116.770131