The proline-rich motif of the proDer p 3 allergen propeptide is crucial for protease-protease interaction.

September 20, 2013 By:
  • Dumez ME
  • Herman J
  • Campisi V
  • Bouaziz A
  • Rosu F
  • Luxen A
  • Vandenberghe I
  • de Pauw E
  • Frere JM
  • Matagne A
  • Chevigne A
  • Galleni M.

The majority of proteases are synthesized in an inactive form, termed zymogen, which consists of a propeptide and a protease domain. The propeptide is commonly involved in the correct folding and specific inhibition of the enzyme. The propeptide of the house dust mite allergen Der p 3, NPILPASPNAT, contains a proline-rich motif (PRM), which is unusual for a trypsin-like protease. By truncating the propeptide or replacing one or all of the prolines in the non-glycosylated zymogen with alanine(s), we demonstrated that the full-length propeptide is not required for correct folding and thermal stability and that the PRM is important for the resistance of proDer p 3 to undesired proteolysis when the protein is expressed in <italic>Pichia pastoris</italic>. Additionally, we followed the maturation time course of proDer p 3 by coupling a quenched-flow assay to mass spectrometry analysis. This approach allowed to monitor the evolution of the different species and to determine the steady-state kinetic parameters for activation of the zymogen by the major allergen Der p 1. This experiment demonstrated that prolines 5 and 8 are crucial for proDer p 3-Der p 1 interaction and for activation of the zymogen.

2013 Sep. PLoS One.8(9):e68014.
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