Gamma-glutamyl carboxylation of vitamin K-dependent proteins is essential for their activation, and a long-term goal of my laboratory is to understand the mechanism and regulation of carboxylation and how dysfunction leads to disease. Carboxylation was originally thought to be important only to hemostasis, but is now known to have a much broader impact on human health, with vitamin K-dependent functions that include calcification, apoptosis, signal transduction and growth control. Carboxylation occurs in the endoplasmic reticulum during the secretion of vitamin K-dependent proteins, which are modified by the combined action of the gamma-glutamyl carboxylase and vitamin K oxidoreductase (VKORC1), as well as unknown components. One area of research is to define this complex process by determining how the secretory process impacts carboxylation, and by identifying other proteins required for carboxylation. Another area of interest is to determine why mutations in the carboxylase cause two distinct diseases, i.e., VKCFD1 in which bleeding is severe or pseudoxanthoma elasticum that is associated with excessive calcification and skin defects. Naturally occurring VKORC1 mutations have also been identified. These mutations alter the response of patients to drugs like warfarin (Coumadin), which is used by millions of people to suppress vitamin K-dependent clotting activity. The mechanism of warfarin inhibition is poorly understood, and a final research focus is to understand warfarin inhibition and the consequence of VKORC1 mutations. Approaches used in our research include mouse models, proteomics, mutagenesis, cellular systems, and biochemical studies.