Figure 2. Mechanism of Required Cofactor Action in Promoting or Inhibiting Blood Clot Formation. Our FRET measurements have revealed that a primary function of the cofactor is to align the enzyme active site at the proper height and orientation above the membrane surface for cleavage of the necessary peptide bond in the substrate. This was demonstrated directly by domain exchange experiments that altered the active site height and/or orientation of the enzyme and thereby eliminated the need for a cofactor to obtain physiological rates of substrate cleavage (Yegneswaran et al., 1999).