Superfolder GFP Is Fluorescent in Oxidizing Environments When Targeted via the Sec Translocon

Traffic. 2011 May;12(5):543-8. doi: 10.1111/j.1600-0854.2011.01168.x. Epub 2011 Feb 25.

Abstract

The ability to study proteins in live cells using genetically encoded fluorescent proteins (FPs) has revolutionized cell biology (1-3). Researchers have created numerous FP biosensors and optimized FPs for specific organisms and subcellular environments in a rainbow of colors (4,5). However, expressing FPs in oxidizing environments such as the eukaryotic endoplasmic reticulum (ER) or the bacterial periplasm can impair folding, thereby preventing fluorescence (6,7). A substantial fraction of enhanced green fluorescent protein (EGFP) oligomerizes to form non-fluorescent mixed disulfides in the ER (6) and EGFP does not fluoresce in the periplasm when targeted via the SecYEG translocon (7). To overcome these obstacles, we exploited the highly efficient folding capability of superfolder GFP (sfGFP) (8). Here, we report sfGFP does not form disulfide-linked oligomers in the ER and maltose-binding protein (MBP) signal sequence (peri)-sfGFP (9) is brightly fluorescent in the periplasm of Escherichia coli. Thus, sfGFP represents an important research tool for studying resident proteins of oxidizing environments.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cell Line
  • Disulfides / chemistry
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism
  • Fluorescence
  • Green Fluorescent Proteins / chemistry*
  • Green Fluorescent Proteins / metabolism*
  • Humans
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Oxidation-Reduction
  • Protein Conformation*
  • Protein Folding*
  • SEC Translocation Channels

Substances

  • Disulfides
  • Escherichia coli Proteins
  • Membrane Proteins
  • SEC Translocation Channels
  • SecE protein, E coli
  • SecG protein, E coli
  • SecY protein, E coli
  • enhanced green fluorescent protein
  • Green Fluorescent Proteins