Elevated serum cholesterol is a major factor in the development of cardiovascular diseases such as coronary heart disease and stroke. Although the initial steps in cholesterol synthesis are well described, the downstream pathway for cholesterol synthesis, from squalene to cholesterol, is relatively poorly characterized, despite its importance to physiology and medicine. My laboratory uses recombinant DNA techniques, enzymology, structural biology and proteomics approaches to characterize squalene monooxygenase, the rate-limiting step in this downstream pathway. Squalene monooxygenase is a 64-kDa flavoprotein bound to the endoplasmic reticulum. Squalene monooxygenase activity is modulated by supernatant protein factor, a 46-kDa cytosolic protein that is thought to serve as a carrier of squalene into and between intracellular membranes. The activity of supernatant protein factor is, in turn, regulated both at the level of expression and, more immediately, through phosphorylation by protein kinases A and C. We are studying the interplay between supernatant protein factor, squalene monooxygenase, protein kinases, and cholesterol levels, in an effort to define the mechanisms that regulate sterol synthesis in the cell.