Directed evolution of APEX2 for electron microscopy and proximity labeling

Nat Methods. 2015 Jan;12(1):51-4. doi: 10.1038/nmeth.3179. Epub 2014 Nov 24.


APEX is an engineered peroxidase that functions as an electron microscopy tag and a promiscuous labeling enzyme for live-cell proteomics. Because limited sensitivity precludes applications requiring low APEX expression, we used yeast-display evolution to improve its catalytic efficiency. APEX2 is far more active in cells, enabling the use of electron microscopy to resolve the submitochondrial localization of calcium uptake regulatory protein MICU1. APEX2 also permits superior enrichment of endogenous mitochondrial and endoplasmic reticulum membrane proteins.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Ascorbate Peroxidases / biosynthesis*
  • Ascorbate Peroxidases / genetics
  • COS Cells
  • Calcium-Binding Proteins / analysis
  • Cation Transport Proteins / analysis
  • Chlorocebus aethiops
  • Directed Molecular Evolution / methods
  • HEK293 Cells
  • HeLa Cells
  • Humans
  • Microscopy, Electron, Transmission / methods*
  • Mitochondria / metabolism
  • Mitochondrial Membrane Transport Proteins / analysis
  • Proteomics / methods*
  • Saccharomyces cerevisiae / enzymology


  • Calcium-Binding Proteins
  • Cation Transport Proteins
  • MICU1 protein, human
  • Mitochondrial Membrane Transport Proteins
  • Ascorbate Peroxidases