METAL IDENTIFICATION IN HUMAN ANTIHEMOPHILIA A-FACTOR (FACTOR-VIII)

Anti-hemophilia A factor (FVIII) consists in different heterodimers of heavy and light chains associated by a metallic ion. The integrity of this complex is indispensable for procoagulant activity. Atomic absor ption spectrometry with Zeeman effect has been applied to determine th e nature of this metal. For this purpose, the different active forms o f FVIII were separated by FPLC and characterized by SDS-PAGE. Two peak s were observed, the first corresponding to different FVIII complexes of high molecular mass (ranging from 210-80 kDa to 110-80 kDa) and the second to the heavy-light chain dimer of 90-80 kDa. In all these acti ve fractions, copper atom was identified and a proportionality was mea sured between the metal concentration and the coagulant activity. Furt hermore, the determination of copper and FVIII concentrations indicate d that only one copper atom is implicated in the 90-80 kDa association .