Protransglutaminase (factor XIII) mediated crosslinking of fibrinogen and fibrin

Plasma factor XIII (plasma protransglutaminase) circulates as an A,B, tetra mer bound to the gamma' variant chains of fibrinogen "2". During clotting t he A subunits of fXIII are cleaved by thrombin to form fXIIIa (transglutami nase) and in the presence of calcium ions, activated A(2)* subunits dissoci ate from the B subunits. When purified plasma fXIII or recombinant cellular factor XIII (,A,) was incubated with fibrinogen in the presence of calcium ions (greater than or equal to 50 muM) a non-synerizing gel formed concomi tant with formation of dimers, followed by Ay polymers, and eventually gamm a trimers and gamma tetramers. As is the case of fXIIIa, the fXIII-mediated crosslinking rate was enhanced in the presence of thiols. After an initial lag period, fXIII catalyzed fibrinogen crosslinking at similar to 75% of t he rate of fXIIIa under typical crosslinking conditions (100 Loewy u/ml, 5 mM CaCl2 & 500 muM DTT). Fibrin was crosslinked about 8 times more rapidly by fXIII than was fibrinogen, and after an initial lag period fXIII crossli nked fibrin at nearly the same rate as fXIIIa. Substituting plasma for puri fied fXIII as the source for fXIII resulted in robust fibrinogen crosslinki ng activity, In contrast to the high level of fXIII-mediated crosslinking a ctivity observed with fibrinogen or fibrin as substrates, when transglutami nation was measured using cadaverine incorporation into casein, fXIII was 3 0-fold less active than fXIIIa. Thus, factor XIII displays constitutive enz ymatic activity with respect to fibrinogen and fibrin. The results further indicate that uncleaved. fXIII in plasma provides a potent source of readil y available crosslinking activity in clotting blood, Fibrinogen 2, whose ga mma 'chains bind fXIII B subunits, was crosslinked 3.5 times more slowly by fXIII than was fibrinogen I (lacking gamma' chains), suggesting that compl ex formation between fibrinogen 2 and plasma fXIII plays a significant role in down-regulating potential plasma fXIII-mediated crosslinking activity. Since fibrin is a considerably better substrate for fXIII than is fibrinoge n, the rate at which crosslinking takes place in a fibrinogen-containing pl asma environment is much lower than it would be if fibrin were present.