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Rhee, Hyun-Woo
Biomimetic Chemistry Lab
Research Interests
  • Site-Specific Protein Labeling in Living Cells

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Proteomic Mapping of the Human Mitochondrial Intermembrane Space in Live Cells via Ratiometric APEX Tagging

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Title
Proteomic Mapping of the Human Mitochondrial Intermembrane Space in Live Cells via Ratiometric APEX Tagging
Author
Hung, VictoriaZou, PengRhee, Hyun-WooUdeshi, Namrata D.Cracan, ValentinSvinkina, TanyaCarr, Steven A.Mootha, Vamsi K.Ting, Alice Y.
Keywords
ENCODED FLUORESCENT REPORTERS; CALCIUM UNIPORTER; LIVING CELLS; CA2+ UPTAKE; MICU1; PROTEINS; BIOLOGY; AUTOPHAGOSOMES; MEMBRANE; COMPLEX
Issue Date
201407
Publisher
CELL PRESS
Citation
MOLECULAR CELL, v.55, no.2, pp.332 - 341
Abstract
Obtaining complete protein inventories for subcellular regions is a challenge that often limits our understanding of cellular function, especially for regions that are impossible to purify and are therefore inaccessible to traditional proteomic analysis. We recently developed a method to map proteomes in living cells with an engineered peroxidase (APEX) that bypasses the need for organellar purification when applied to membrane-bound compartments; however, it was insufficiently specific when applied to unbounded regions that allow APEX-generated radicals to escape. Here, we combine APEX technology with a SILAC-based ratiometric tagging strategy to substantially reduce unwanted background and achieve nanometer spatial resolution. This is applied to map the proteome of the mitochondrial intermembrane space (IMS), which can freely exchange small molecules with the cytosol. Our IMS proteome of 127 proteins has >94% specificity and includes nine newly discovered mitochondrial proteins. This approach will enable scientists to map proteomes of cellular regions that were previously inaccessible.
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DOI
http://dx.doi.org/10.1016/j.molcel.2014.06.003
ISSN
1097-2765
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