Folding and stability of the leucine-rich repeat domain of internalin B from Listeria monocytogenes

Alexander Freiberg, Matthias P. Machner, Wolfgang Pfeil, Wolf Dieter Schubert, Dirk W. Heinz, Robert Seckler

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24 Scopus citations


Internalin B (InlB), a surface protein of the human pathogen Listeria monocytogenes, promotes invasion into various host cell types by inducing phagocytosis of the entire bacterium. The N-terminal half of InlB (residues 36-321, InlB321), which is sufficient for this process, contains a central leucine-rich repeat (LRR) domain that is flanked by a small α-helical cap and an immunoglobulin (Ig)-like domain. Here we investigated the spectroscopic properties, stability and folding of InlB 321 and of a shorter variant lacking the Ig-like domain (InlB 248). The circular dichroism spectra of both protein variants in the far ultraviolet region are very similar, with a characteristic minimum found at ~200 nm, possibly resulting from the high 310-helical content in the LRR domain. Upon addition of chemical denaturants, both variants unfold in single transitions with unusually high cooperativity that are fully reversible and best described by two-state equilibria. The free energies of GdmCl-induced unfolding determined from transitions at 20°C are 9.9(±0.8)kcal/mol for InlB321 and 5.4(±0.4)kcal/mol for InlB248. InlB321 is also more stable against thermal denaturation, as observed by scanning calorimetry. This suggests, that the Ig-like domain, which presumably does not directly interact with the host cell receptor during bacterial invasion, plays a critical role for the in vivo stability of InlB.

Original languageEnglish (US)
Pages (from-to)453-461
Number of pages9
JournalJournal of Molecular Biology
Issue number2
StatePublished - Mar 19 2004
Externally publishedYes


  • 3 -helix
  • CD, circular dichroism
  • DSC, differential scanning calorimetry
  • GdmCl, guanidinium chloride
  • Leucine-rich repeat
  • Protein folding
  • Protein stability
  • Spectroscopy
  • [GdmCl], midpoint of a GdmCl-induced unfolding transition

ASJC Scopus subject areas

  • Molecular Biology
  • Biophysics
  • Structural Biology


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