Heparin promotes the formation of 1° FXaα-ATIII and FXaβ-ATIII complexes from the free enzymes and antithrombin III. It further stimulates the transformations of the 1° complexes into the corresponding 3° complexes. Additionally, it stimulates the degradation of the 1° FXaα-ATIII and FXaβ-ATIII complexes into free FXaα, FXaβ and ATIIIM. Protamine sulfate (PS) stimulates the transformation of FXaα into FXaβ and hence to FXaγ. It also stimulates the transformation of the 1°-, 2°-, and 3°- FXα-ATIII complexes into their corresponding β-complexes. It further promotes the degradation of the 1° FXaα- and FXaβ-ATIII complexes into their corresponding 3°- FXaα- and FXaβ-ATIII complexes, with the concommittant release of FXaγ. The addition of PS to FXa/ATIII/H mixtures results in a reduction in free FXa, ATIII, and 1° Xa-ATIII complex formation, together with the concommittant increase in 3°-FXa- ATIII complex formation and release of FXaγ. The likeliest explanation of these results resides in the removal of the effective free heparin as a consequence of the generation of a stable heparin/PS salt upon the addition of the PS to the FXa/ATIII/H mixtures, thereby effectively lowering clotting times. 1. Introduction Factor X is located at the juncture of the intrinsic and extrinsic coagulation cascade pathways. Factor X has two active forms (FXaα and FXaβ) that differ in molecular weight by about 4000 daltons. Factor Xaβ can be degradatively inactivated (autolyzed) to form FXaγ upon cleavage at the –NH2 terminal of the active site [1–3]. The serpin antithrombin III (ATIII) is the primary naturally occurring inhibitor of Factor Xa. Factor Xa and antithrombin III form a 1？:？1 molar complex via covalent bonding [3–5]. Upon hydrolysis of this bond, a modified form of ATIII is produced (ATIIIM). ATIIIM runs at a slightly higher apparent molecular weight than native ATIII in SDS-PAGE [3, 5, 6]. The polysulfonated glycosaminoglycan heparin (H) is administered clinically as an anticoagulant. H acts to catalyze interactions of ATIII with factors Xa and IIa, among other coagulation factors. Heparin fractions have varying affinity for ATIII, with the smallest fraction of high affinity being the pentasaccharide. Administration of H increases the FXa-ATIII complex formation by at least an order of magnitude [7] or several hundredfold when also in the presence of physiological levels of Ca2+[8–10]. Further, ATIIIM increases 20–30% in the presence of FXa and high-affinity heparin [6, 11]. Positively charged protamine sulfate (PS) has long been used clinically to neutralize heparin
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