Related Experiment Videos
Human factor VIII: purification from commercial factor VIII concentrate, characterization, identification and
Biochimica Et Biophysica Acta
|October 17, 1986
Summary
Researchers purified human factor VIII, revealing its complex polypeptide structure and activation by thrombin and factor Xa. Activated protein C inactivated factor VIII, providing insights into coagulation factor regulation.
Area of Science:
- Biochemistry
- Hematology
Background:
- Factor VIII is a critical protein in the blood coagulation cascade.
- Understanding its structure and activation is vital for treating bleeding disorders.
Purpose of the Study:
- To purify and characterize human factor VIII.
- To investigate the activation and inactivation mechanisms of factor VIII.
Main Methods:
- Purification of factor VIII from commercial concentrate.
- Analysis of purified factor VIII using SDS-PAGE and Sephadex G200 gel filtration.
- Investigating factor VIII activation with thrombin, factor Xa, and activated protein C.
- Radioiodination of factor VIII for further characterization.
Main Results:
- Purified factor VIII yielded 12% with a specific activity of 8,000 units/mg.
- SDS-PAGE revealed polypeptides ranging from Mr 80,000 to Mr 208,000; Sephadex G200 showed an apparent molecular weight of 270,000.
- Thrombin activation led to the formation of an Mr 92,000 polypeptide, while factor Xa also activated factor VIII.
- Activated protein C inactivated factor VIII's coagulant activity.
- Radioiodinated factor VIII exhibited identical characteristics to unlabeled factor VIII and could be precipitated by anti-factor VIII antibodies.
Conclusions:
- Human factor VIII is a complex protein composed of multiple polypeptides.
- Factor VIII activation involves specific proteolytic events.
- Activated protein C plays an inhibitory role in factor VIII function.