Osteocalcin binds tightly to the gamma-glutamylcarboxylase at a site distinct from that of the other known vitamin K-dependent proteins

Vitamin K-dependent proteins contain a propeptide that is required for reco gnition by the enzyme gamma-glutamylcarboxylase. Substrates used in vitro f or carboxylation studies lacking a prosequence are characterized by K-m val ues in the millimolar range, whereas the K-m for peptides containing a pros equence is three or four orders of magnitude smaller. Here we report that d escarboxy-osteocalcin is an exception in this respect. With descarboxy-oste ocalcin in purified propeptide-free recombinant carboxylase, the K-m was 1. 8 mu M. Furthermore, osteocalcin was an inhibitor of descarboxy-osteocalcin carboxylation with a K-i of 76 mu M. In contrast with the other vitamin K- dependent proteins, free propeptides do not inhibit descarboxy-osteocalcin carboxylation. Moreover, propeptide-containing substrates were inhibited ne ither by osteocalcin nor by its propeptide. From our studies we conclude th at descarboxy-osteocalcin must have an internal recognition sequence that b inds to gamma-glutamylcarboxylase at a site different from the propeptide-r ecognition site.