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目的研究FGFC1对各纤溶因子构象的影响及FGFC1促进纤溶反应的机制。方法采用圆二色谱法研究FGFC1对各纤溶因子结构的影响,采用发色底物法研究FGFC1对各纤溶因子活性的影响,采用SDSPAGE电泳法研究FGFC1在纤溶酶原和单链尿激酶型纤溶酶原激活剂构成的相互活化反应体系中的作用。结果远紫外区的CD结果表明FGFC1在23~115μmol·L-1的浓度范围内均使各纤溶因子的二级结构发生变化,这变化都导致酶分子柔性增加、更易发生反应;近紫外区的CD结果表明,FGFC1使纤溶酶原在285nm处的摩尔旋光度增大。CD研究结果表明FGFC1对单链尿激酶型纤溶酶原激活剂和尿激酶型纤溶酶原激活剂结构的影响较为微弱,对纤溶酶及纤溶酶原结构的影响复杂而深刻。运用发色底物法检测加入不同浓度FGFC1后各纤溶因子的酶活力,排除了FGFC1对纤溶因子的结构影响而导致的纤溶因子失活的可能性。在体外构筑的纤溶反应体系中,SDS-PAGE的结果显示FGFC1的加入促进FITC-纤维蛋白原的降解,表明FGFC1加速纤溶酶原和单链尿激酶型纤溶酶原激活剂的相互活化作用。结论 FGFC1作用于纤溶酶原并辅以改变单链尿激酶型纤溶酶原激活剂的二级结构发挥纤溶促进作用。
Aim To investigate the effect of FGFC1 on the conformation of fibrinolytic factor and the mechanism of FGFC1 on fibrinolysis. Methods The effects of FGFC1 on the structure of fibrinolytic factors were studied by circular dichroism spectroscopy. The effect of FGFC1 on the activity of fibrinolytic factors was investigated by chromogenic substrate method. The effect of FGFC1 on the activities of fibrinolytic enzymes and single-chain urokinase Type plasminogen activator composed of mutual activation reaction system role. Results The results of CD in far ultraviolet region showed that FGFC1 changed the secondary structure of fibrinolytic factors in the concentration range of 23 ~ 115μmol·L-1, which led to the increase of the flexibility of the enzyme molecules and the more reaction. The CD results show that FGFC1 increases the molar rotation of plasminogen at 285 nm. CD results show that FGFC1 on single-chain urokinase-type plasminogen activator and urokinase-type plasminogen activator structure is relatively weak, the plasmin and plasminogen structure complex and profound. The chromogenic substrate method was used to detect the enzyme activity of each fibrinolytic factor after adding different concentrations of FGFC1, and the possibility of inactivation of fibrinolytic factors was eliminated due to the structural influence of FGFC1 on fibrinolytic factors. In the fibrinolytic system constructed in vitro, the results of SDS-PAGE showed that the addition of FGFC1 promoted the degradation of FITC-fibrinogen, indicating that FGFC1 accelerated the mutual activation of plasminogen and single-chain urokinase-type plasminogen activator effect. Conclusion FGFC1 acts on plasminogen and supplements to change the secondary structure of single-chain urokinase-type plasminogen activator to exert fibrinolysis promoting effect.