THE ROLE OF ANNEXIN-II TETRAMER IN THE ACTIVATION OF PLASMINOGEN

Annexin II tetramer (AIIt) is a major Ca2+-binding protein of endothel ial cells which has been shown to exist on both the intracellular and extracellular surfaces of the plasma membrane. In this report, Eve dem onstrate that AIIt stimulates the activation of plasminogen by facilit ating the tissue plasminogen activator (t-PA)-dependent conversion of plasminogen to plasmin, Fluid-phase AIIt stimulated the rate of activa tion of [Glu]plasminogen about 341-fold compared with an approximate 6 -fold stimulation by annexin II. AIIt bound to [Glu]plasminogen(S741C- fluorescein) with a K-d of 1.26 +/- 0.04 mu M (mean +/- S.D., n = 3) a nd this interaction resulted in a large conformational change in [Glu] plasminogen. Kinetic analysis established that AIIt produces a large i ncrease of about 190-fold in the k(cat, app) and a small increase in t he K-m,K- app which resulted in a 90-fold increase in the catalytic ef ficiency (k(cat)/K-m) of t-PA for [Glu]plasminogen. AIIt also stimulat ed the t-PA-dependent activation of [Lys]plasminogen about 28-fold. Fu rthermore, other annexins such as annexin I, V, or VI did not produce comparable activation of t-PA-dependent conversion of [Glu]plasminogen to plasmin, The stimulation of the activation of [Glu]plasminogen by AIIt was Ca2+-independent and inhibited by epsilon-aminocaproic acid. AIIt bound to human 293 cells potentiated t-PA-dependent plasminogen a ctivation, AIIt that was bound to phospholipid vesicles or heparin als o stimulated the activation of [Glu]plasminogen 5- or 11-fold, respect ively. Furthermore, immunofluorescence labeling of nonpermeabilized HU VEC revealed a punctated distribution of AIIt subunits on the cell sur face, These results therefore identify AIIt as a potent in vitro activ ator of plasminogen.