The porposed research program will continue to investigate the role of proteins containing the vitamin K-dependent, calcium binding, amino acid, gammcarboxyglutamic acid (Gla) in calcific cardiovascular disease. Background work has established Gla to occur at the calcium binding sites of the vitamin K dependent coagulation factors and the bone protein, osteocalcin, via a vitamin K-dependent enzymatic, post-translational, carboxylation of specific glutamic acid residues. In the case of the current program, Gla containing proteins have been demonstrated to occur in pathologic calcifications involving atherosclerotic plaque, calcific aortic stenosis, and porcine xenograft valve calcification. Characterization of the Gla containing proteins isolated from calcified atherosclerotic plaque has resulted in the isolation and characterization of a unique Gla containing protein of 80000 modelcular weight known as atherocalcin; osteocalcin is the only other identifiable Gla-containing protein thus far. Work to data also indicates that Gla content correlates with the severity of pathologic calcification; trace levels of Gla are detectable in apparently normal tissue. Finally, an experimental model of porcine xenograft valve calcification has been developed for the use in studying the role of Gla containing proteins in cardiovascular calcifications. The specific aims for the continuing program are: 1) to continue to characterize the Gla containing proteins of ahterosclerotic plaque; 2) to pursue further studies into the role of Gla containing proteins in cardiovascular diseas via biochemical studies of pathologic material; and 3) to study experimental valvar calcifications, in order to explore the cellular and molecular determinants of the Gla-protein-calcium interaction in developing lesions. The ultimate goal of the program is to attain a sufficient understanding of this problem so that improved appoachs to therapy and prevention of calcific cardiovascular disease may be possible.