Abstract
Coagulation factor XI is activated by thrombin or factor XIIa resulting in a conformational
change that converts the catalytic domain into its active form and exposing exosites
for factor IX on the apple domains. Although crystal structures of the zymogen factor
XI and the catalytic domain of the protease are available, the structure of the apple
domains and hence the interactions with the catalytic domain in factor XIa are unknown.
We now used chemical footprinting to identify lysine residue containing regions that
undergo a conformational change following activation of factor XI. To this end, we
employed tandem mass tag in conjunction with mass spectrometry. Fifty-two unique peptides
were identified, covering 37 of the 41 lysine residues present in factor XI. Two identified
lysine residues that showed altered flexibility upon activation were mutated to study
their contribution in factor XI stability or enzymatic activity. Lys357, part of the
connecting loop between A4 and the catalytic domain, was more reactive in factor XIa
but mutation of this lysine residue did not impact on factor XIa activity. Lys516
and its possible interactor Glu380 are located in the catalytic domain and are covered
by the activation loop of factor XIa. Mutating Glu380 enhanced Arg369 cleavage and
thrombin generation in plasma. In conclusion, we have identified novel regions that
undergo a conformational change following activation. This information improves knowledge
about factor XI and will contribute to development of novel inhibitors or activators
for this coagulation protein.
Keywords
factor XI - conformational change - chemical footprinting - mass spectrometry
