Abstract
Tail-anchored proteins are a group of membrane proteins oriented with their amino terminus in the cytoplasm and their carboxy terminus embedded in intracellular membranes. This group includes the apoptosis-mediating proteins of the Bcl-2 family as well as the vesicle targeting proteins of the SNARE group, among others. A stretch of hydrophobic amino acids at the extreme carboxy terminus of these proteins serves both as a membrane anchor and as a targeting signal. Tail-anchored proteins are differentially targeted to either the endoplasmic reticulum or the mitochondrial outer membrane and the mechanism which accomplishes this selective targeting is poorly understood. Here we define important characteristics of the signal/anchor region which directs proteins to the mitochondrial outer membrane. We have created an artificial sequence consisting of a stretch of 16 leucines bounded by positively charged amino acids. Using this template we demonstrate that moderate hydrophobicity distinguishes the mitochondrial tail-anchor sequence from that of the endoplasmic reticulum tail-anchor sequence. A change as small as introduction of a single polar residue into a sequence that otherwise targets to the endoplasmic reticulum can substantially switch targeting to the mitochondrial outer membrane. Further we show that a mitochondrially targeted tail-anchor has a higher propensity for the formation of alpha-helical structure than a sequence directing tail-anchored proteins to the endoplasmic reticulum.
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Abbreviations
- SNARE:
-
Soluble N-ethylmaleimide factor receptor
- ER:
-
Endoplasmic reticulum
- VAMP:
-
Vesicle associated membrane protein
- MOA:
-
Monoamine oxidase A
- GFP:
-
Green fluorescent protein
- DPC:
-
Dodecylphosphocholine
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Acknowledgements
This work was supported by the Brown Cancer Center, Louisville, Kentucky, the National Institutes of Health (COBRE 5P20RR018733-03), and the American Heart Association (Pre-doctoral grant to SB). We also gratefully acknowledge the invaluable help of Dr. Brad Chaires and Dr. Nichola Garbett for their expert advice and for use of the Brown Cancer Center Biophysics core for circular dichroism measurements. The technical assistance of Ms. Helen Lynn, Ms. Lori Davies, and Mr. Angelo Wilson in early studies is gratefully acknowledged.
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Wattenberg, B.W., Clark, D. & Brock, S. An Artificial Mitochondrial Tail Signal/Anchor Sequence Confirms a Requirement for Moderate Hydrophobicity for Targeting. Biosci Rep 27, 385–401 (2007). https://doi.org/10.1007/s10540-007-9061-0
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DOI: https://doi.org/10.1007/s10540-007-9061-0