A site on factor XII required for productive interactions with polyphosphate.

Area of Science:

• Biochemistry
• Hematology
• Molecular Biology

Background:

• Plasma contact activation involves factor XII (FXII) binding to surfaces and conversion to FXIIa.
• FXIIa activates prekallikrein and factor XI (FXI).
• The FXII first epidermal growth factor-1 (EGF1) domain is critical for polyphosphate-dependent FXII activity.

Purpose of the Study:

• To identify specific amino acids within the FXII EGF1 domain essential for polyphosphate-mediated FXII functions.
• To elucidate the role of these amino acids in FXII activation and downstream protease activation.

Main Methods:

• Alanine substitutions were introduced into basic residues within the FXII EGF1 domain.
• Wild-type FXII and a variant with a Pro-HGFA EGF1 domain served as controls.
• Proteins were assessed for activation, prekallikrein and FXI activation, polyphosphate binding, plasma clotting, and thrombosis in a mouse model.

Main Results:

• FXII variants with alanine substitutions at Lys73, Lys74, Lys76, or Lys76, His78, Lys81 showed impaired activation with polyphosphate.
• These variants exhibited significantly reduced FXII activity in clotting assays and diminished polyphosphate binding.
• FXIIa variants demonstrated defects in surface-dependent FXI activation and poor reconstitution in a thrombosis model.

Conclusions:

• Lysine residues 73, 74, 76, and 81 in the FXII EGF1 domain form a polyphosphate binding site.
• This binding site is indispensable for surface-dependent FXII activation and its physiological functions in hemostasis and thrombosis.