The first two steps of the steroidogenic pathway were reproduced in Saccharomyces cerevisiae. Engineering of sterol biosynthesis by disruption of the 22-desaturase gene and introduction of the Arabidopsis thaliana 7-reductase activity and coexpression of bovine side chain cleavage cytochrome P450, adrenodoxin, and adrenodoxin reductase, lead to pregnenolone biosynthesis from a simple carbon source. Following additional coexpression of human 3-beta-hydroxysteroid dehydrogenase/isomerase, pregnenolone is further metabolized to progesterone. Steroid formation appears to be coupled to yeast sterol biosynthesis.