The Proline-Rich Motif of the proDer p 3 Allergen Propeptide Is Crucial for Protease-Protease Interaction

Marie Eve Dumez, Julie Herman, Vincenzo Campisi, Ahlem Bouaziz, Frédéric Rosu, André Luxen, Isabel Vandenberghe, Edwin de Pauw, Jean Marie Frère, André Matagne, Andy Chevigné, Moreno Galleni*

*Corresponding author for this work

    Research output: Contribution to journalArticleResearchpeer-review

    4 Citations (Scopus)

    Abstract

    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 Pichia pastoris. 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.

    Original languageEnglish
    Article numbere68014
    JournalPLoS ONE
    Volume8
    Issue number9
    DOIs
    Publication statusPublished - 20 Sep 2013

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