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Published May 7, 2013 | Published + Supplemental Material
Journal Article Open

Precise timing of ATPase activation drives targeting of tail-anchored proteins


The localization of tail-anchored (TA) proteins, whose transmembrane domain resides at the extreme C terminus, presents major challenges to cellular protein targeting machineries. In eukaryotic cells, the highly conserved ATPase, guided entry of tail-anchored protein 3 (Get3), coordinates the delivery of TA proteins to the endoplasmic reticulum. How Get3 uses its ATPase cycle to drive this fundamental process remains unclear. Here, we establish a quantitative framework for the Get3 ATPase cycle and show that ATP specifically induces multiple conformational changes in Get3 that culminate in its ATPase activation through tetramerization. Further, upstream and downstream components actively regulate the Get3 ATPase cycle to ensure the precise timing of ATP hydrolysis in the pathway: the Get4/5 TA loading complex locks Get3 in the ATP-bound state and primes it for TA protein capture, whereas the TA substrate induces tetramerization of Get3 and activates its ATPase reaction 100-fold. Our results establish a precise model for how Get3 harnesses the energy from ATP to drive the membrane localization of TA proteins and illustrate how dimerization-activated nucleotide hydrolases regulate diverse cellular processes.

Additional Information

© 2013 National Academy of Sciences. Edited by Douglas C. Rees, Howard Hughes Medical Institute, Caltech, Pasadena, CA, and approved March 26, 2013 (received for review December 19, 2012). Published online before print April 22, 2013. We thank J. Chartron, H. Gristick, and C. J. M. Suloway for expression constructs, purification protocols, and critical discussions; M. Sachs and C. Wu for help with yeast translation extracts; R. Schekman for help with yeast microsomes; and D. C. Rees and members of the S.-o.S. and W.M.C. groups for helpful comments. This work was supported by career awards from the David and Lucile Packard Foundation and the Henry Dreyfus Foundation (to S.-o.S.), National Science Foundation Graduate Research Fellowship DGE-1144469 (to M.E.R.), National Institutes of Health (NIH) Training Grant 5T32GM007616-33 (to M.R.), and NIH Grant R01 GM097572 (to W.M.C.). Author contributions: M.E.R., M.R., and S.-o.S. designed research; M.E.R. and M.R. performed research; W.M.C. contributed new reagents/analytic tools; M.E.R., M.R., and S.-o.S. analyzed data; and M.E.R., M.R., and S.-o.S. wrote the paper.

Attached Files

Published - PNAS-2013-Rome-7666-71.pdf

Supplemental Material - pnas.201222054SI.pdf


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August 22, 2023
August 22, 2023