We determined the genomic structure of the human gene encoding 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, which catalyzes the conversion of HMG-CoA to mevalonate and is the rate-limiting and major regulatory enzyme in sterol biosynthesis. The gene is more than 21 kb long, about five times the size of its corresponding cDNA. It consists of 20 exons, ranging in size from 68 to 1809bp. An amino-terminal hydrophobic membrane-bound domain is encoded by exons 2-10, a flexible linker domain by exons 10 and 11, and the catalytic domain by exons 11-20. Exons 3-7 encode a sterol-sensing domain. We compared its genomic structure in this region with the sterol-sensing domains of three related genes, sterol regulatory element binding protein (SREBP) cleavage-activating protein (SCAP), Niemann-Pick type C1 protein (NPC1), and a morphogen receptor, Patched. Two of the five positions of introns in the sterol-sensing domain of the HMG-CoA reductase gene were identical to the exon/intron organization of this domain in the related human genes, but these positions of introns were not conserved in homologues from lower organisms, except in one instance. The data suggested that exon-shuffling may have occurred during relatively recent evolution: this would account for the structural similarity of this domain in four quite different human proteins.