The crystalloid endoplasmic reticulum (ER) of UT-1 cells is a specialized smooth ER that houses 3-hydroxy-3-methylglutaryl-CoA reductase, a membrane protein that regulates endogenous cholesterol synthesis. The biogenesis of this ER is coupled to the over production of 3-hydroxy-3-methylglutaryl-CoA reductase. To understand better this membrane system and the relationship between the synthesis of a membrane protein and the formation of membrane, we have purified the crystalloid ER. Purified crystalloid ER did not contain significant amounts of membrane derived from the Golgi apparatus, mitochondria, or plasma membrane. Approximately 24% of the protein in this organelle corresponded to 3-hydroxy-3-methylglutaryl-CoA reductase; however, at least eight other proteins were detected by gel electrophoresis. One of these proteins (Mr 73,000) was as abundant as reductase. These results suggest that the biogenesis of this ER involves the coordinate synthesis of multiple membrane and content proteins.