The A1 and A2 subunits of factor VIIIa synergistically stimulate factor IXa catalytic activity

被引:34
作者
Fay, PJ
Koshibu, K
Mastri, M
机构
[1] Univ Rochester, Med Ctr, Vasc Med Unit, Sch Med & Dent,Dept Med, Rochester, NY 14642 USA
[2] Univ Rochester, Sch Med & Dent, Dept Biochem & Biophys, Rochester, NY 14642 USA
关键词
D O I
10.1074/jbc.274.22.15401
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Factor VIIIa, the protein cofactor for factor IXa, is comprised of A1, A2, and A3-C1-C2 subunits. Recently, we showed that isolated A2 subunit enhanced the k(cat) for factor IXa-catalyzed activation of factor X by similar to 100-fold (similar to 1 min(-1)), whereas isolated A1 or A3-C1-C2 subunits showed no effect on this rate (Fay, P. J., and Koshibu, K. J. (1998) J, Biol, Chem, 273, 19049-19054), However, A1 subunit increased the A2-dependent stimulation by similar to 10-fold. The K-m for factor X in the presence of A2 subunit was unaffected by A1 subunit, whereas the k(cat) observed in the presence of saturating A1 and A2 subunits (similar to 15 min(-1)) represented 5-10% of the value observed for native factor VIIIa (similar to 200 min(-1)). An anti-A1 subunit antibody that blocks the association of A2 eliminated the A1-dependent contribution to factor IXa activity, Inclusion of both A1 and A2 subunits resulted in greater increases in the fluorescence anisotropy of fluorescein-Phe-Phe-Arg factor IXa than that observed for A2 subunit alone and approached values obtained with factor VIIIa, These results indicate that A1 subunit alters the A2 subunit-dependent modulation of the active site of factor IXa to synergistically increase cofactor activity, yielding an overall increase in k(cat) of over 1000-fold compared with factor IXa alone.
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页码:15401 / 15406
页数:6
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