TRANSGLUTAMINASE-CATALYZED CROSS-LINKING OF OSTEOPONTIN IS INHIBITED BY OSTEOCALCIN

Osteocalcin, the most abundant noncollagenous protein of bone matrix, has been demonstrated to inhibit bone growth by gene knockout experime nts (Ducy, P., Desbois, C., Boyce, B., Pinero, G., Story, B., Dunstan, C., Smith, E., Bonadio, J., Goldstein, S., Gundberg, C., Bradley, A., and Karsenty, G. (1996) Nature 382, 448-452). Its specific functional mechanism in bone metabolism is, however, largely unknown. In this st udy, we provide evidence that osteocalcin has an inhibitory effect on tissue transglutaminase activity, as measured by cross-linking of oste opontin, another bone matrix protein. Using a set of synthetic peptide s, we found that the inhibitory activity resided within the first 13 N -terminal amino acid residues of osteocalcin. An N-terminal peptide al so inhibited cross-linking of another tissue transglutaminase substrat e, beta-casein. The inhibitory peptide was shown to have affinity for the substrates of transglutaminase rather than for the enzyme. Since t he N terminus of osteocalcin exhibits homology to the substrate recogn ition site sequences of two transglutaminases, we conclude that the in hibitory effect is most likely due to competition with the enzyme for the transglutaminase-binding region of the substrates, osteopontin and beta-casein, which prevents access of the enzyme to them to perform i ts function. The interference of osteocalcin with osteopontin cross-li nking gives osteocalcin a new potential function as the first protein inhibitor of tissue transglutaminase. This suggests a specific role an d a plausible mechanism for it as a modulator of maturation, stabiliza tion, and calcification of bone matrix.