Application of split-green fluorescent protein for topology mapping membrane proteins in Escherichia coli

Stephen Toddo, Bill Soderstrom, Isolde Palombo, Gunnar von Heijne, Morten Nørholm, Daniel O. Daley

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A topology map of a membrane protein defines the location of transmembrane helices and the orientation of soluble domains relative to the membrane. In the absence of a high-resolution structure, a topology map is an essential guide for studying structurefunction relationships. Although these maps can be predicted directly from amino acid sequence, the predictions are more accurate if combined with experimental data, which are usually obtained by fusing a reporter protein to the C-terminus of the protein. However, as reporter proteins are large, they cannot be used to report on the cytoplasmic/periplasmic location of the N-terminus of a protein. Here, we show that the bimolecular split-green fluorescent protein complementation system can overcome this limitation and can be used to determine the location of both the N- and C-termini of inner membrane proteins in Escherichia coli.
Original languageEnglish
JournalProtein Science
Issue number10
Pages (from-to)1571-1576
Publication statusPublished - 2012


  • structure-function relationship
  • Facultatively Anaerobic Gram-Negative Rods Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Enterobacteriaceae [06702] Escherichia coli species strain-BL21(DE3)
  • inner membrane protein N-terminus, C-terminus
  • split-green fluorescent protein complementation system
  • 10060, Biochemistry studies - General
  • 31000, Physiology and biochemistry of bacteria
  • topology mapping laboratory techniques
  • Biochemistry and Molecular Biophysics


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