Serine protease activation is typically controlled by proteolytic cleavage of the scissile bond, resulting in spontaneous formation of the activating Ile(16)-Asp(194) salt bridge. The initiating coagulation protease factor VIIa (VIIa) differs by remaining in a zymogen-like conformation that confers the control of catalytic activity to the obligatory cofactor and receptor tissue factor (TF). This study demonstrates that the unusual hydrophobic Met(156) residue contributes to the propensity of the VIIa protease domain to remain in a zymogen-like conformation. Mutation of Met(156) to Gln, which is found in the same position of the highly homologous factor IX, had no influence on the amidolytic and proteolytic activity of TF-bound VIIa. Furthermore, the mutation did not appreciably stabilize the labile Ile(16)-Asp(194) salt bridge in the absence of cofactor. VIIa(Gln156) had increased affinity for TF, consistent with a long range conformational effect that stabilized the cofactor binding site in the VIIa protease domain. Notably, in the absence of cofactor, amidolytic and proteolytic function of VIIa(Gln156) were enhanced 3- and 9-fold, respectively, compared with wild-type VIIa. The mutation thus selectively influenced the catalytic activity of free VIIa, identifying the Met(156) residue position as a determinant for the zymogen-like properties of free VIIa.