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Hydrogen/Deuterium Exchange Mass Spectrometry Reveals Specific Changes in the Local Flexibility of Plasminogen Activator Inhibitor 1 upon Binding to the Somatomedin B Domain of Vitronectin

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  • Morten Beck Trelle, Institut for Biokemi og Molekylær Biologi, Danmark
  • Daniel Hirschberg, Institut for Biokemi og Molekylær Biologi, Danmark
  • Anna Jansson, Institut for Biokemi og Molekylær Biologi, Danmark
  • Michael Ploug, Danmark
  • Peter Roepstorff, Institut for Biokemi og Molekylær Biologi, Danmark
  • Peter Andreasen, Danmark
  • Thomas Jørgensen, Aalborg Universitet, Danmark
The native fold of plasminogen activator inhibitor 1 (PAI-1) represents an active metastable conformation that spontaneously converts to an inactive latent form. Binding of the somatomedin B domain (SMB) of the endogenous cofactor vitronectin to PAI-1 delays the transition to the latent state and increases the thermal stability of the protein dramatically. We have used hydrogen/deuterium exchange mass spectrometry to assess the inherent structural flexibility of PAI-1 and to monitor the changes induced by SMB binding. Our data show that the PAI-1 core consisting of β-sheet B is rather protected against exchange with the solvent, while the remainder of the molecule is more dynamic. SMB binding causes a pronounced and widespread stabilization of PAI-1 that is not confined to the binding interface with SMB. We further explored the local structural flexibility in a mutationally stabilized PAI-1 variant (14-1B) as well as the effect of stabilizing antibody Mab-1 on wild-type PAI-1. The three modes of stabilizing PAI-1 (SMB, Mab-1, and the mutations in 14-1B) all cause a delayed latency transition, and this effect was accompanied by unique signatures on the flexibility of PAI-1. Reduced flexibility in the region around helices B, C, and I was seen in all three cases, which suggests an involvement of this region in mediating structural flexibility necessary for the latency transition. These data therefore add considerable depth to our current understanding of the local structural flexibility in PAI-1 and provide novel indications of regions that may affect the functional stability of PAI-1.
OriginalsprogEngelsk
TidsskriftBiochemistry
Vol/bind51
Nummer41
Sider (fra-til)8256–8266
Antal sider11
ISSN0006-2960
DOI
StatusUdgivet - 2012

    Forskningsområder

  • Serpins; mass spectrometry;

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