PARTIAL IDENTIFICATION OF THE ZN2-BINDING SITES IN FACTOR-XII AND ITSACTIVATION DERIVATIVES()

With the purpose of identifying the Zn2+-binding sites in factor XII, the effect of chemical modification of His, Glu and Asp residues, amin o acids known to participate in the catalytic coordination binding of Zn2+ in a number of Zn2+-binding proteins, was analysed. The number of modifiable His residues in factor XII and alpha-factor XIIa was 16.0/-0.7 and 17.3+/-0.7, respectively. When factor XII/alpha-factor XIIa was incubated with saturating concentrations of Zn2+ before the diethy lpyrocarbonate modification of the His residues, these numbers were re duced to 6.3+/-0.1 and 8.2+/-0.5, indicating that ten and nine His res idues, respectively are involved in the binding. Analysis of the Zn2+- binding capacity of factor XII, alpha-factor XIIa and beta-factor XIIa showed that while factor XII contains four Zn2+-binding sites, alpha- factor XIIa had only three and beta-factor XIIa had none. Modification of the His residues resulted in a complete loss of Zn2+-binding while Asp/Glu modification resulted in loss of two and one Zn2+-binding sit es in factor XII and alpha-factor XIIa, respectively. This suggests th at two of the four sites in factor XII contain His residues, exclusive ly, while the two others are comprised of two His residues and one Asp /Glu residue. One of the latter is lost when factor XII is activated t o alpha-factor XIIa. Two of the sites are suggested to be located at p ositions His40-His44 and His78-His82. The location of the remaining tw o sites are reduced to four possible positions.