The propeptides of the vitamin K-dependent proteins possess different affinities for the vitamin K-dependent carboxylase

The vitamin K-dependent gamma-glutamyl carboxylase catalyzes the modificati on of specific glutamates in a number of proteins required for blood coagul ation and associated with bone and calcium homeostasis. All known vitamin K -dependent proteins possess a conserved eighteen-amino acid propeptide sequ ence that is the primary binding site for the carboxylase, We compared the relative affinities of synthetic propeptides of nine human vitamin R-depend ent proteins by determining the inhibition constants (K-i) toward a factor IX propeptide/gamma-carboxyglutamic acid domain substrate. The Ki values fo r six of the propeptides (factor X, matrix Gla protein, factor VII, factor IX, PRGP1, and protein S) were between 2-35 nM, with the factor X propeptid e having the tightest affinity. In contrast, the inhibition constants for t he propeptides of prothrombin and protein C are similar to 100-fold weaker than the factor X propeptide, The propeptide of bone Gla protein demonstrat es severely impaired carboxylase binding with an inhibition constant of at least 200,000-fold weaker than the factor X propeptide, This study demonstr ates that the affinities of the propeptides of the vitamin K-dependent prot eins vary over a considerable range; this may have important physiological consequences in the levels of vitamin K-dependent proteins and the biochemi cal mechanism by which these substrates are modified by the carboxylase.