Betulinic acid is a plant-based triterpenoid that has been recognized for its antitumor and anti-HIV activities. The level of betulinic acid in its natural hosts is extremely low. In the present study, we constructed betulinic acid biosynthetic pathway in Saccharomyces cerevisiae by metabolic engineering. Given the betulinic acid forming pathways sharing the common substrate acetyl-CoA with fatty acid synthesis, the metabolic fluxes between the two pathways were varied by changing gene expressions, and their effects on betulinic acid production were investigated. We constructed nine S. cerevisiae strains representing nine combinations of the flux distributions between betulinic acid and fatty acid pathways. Our results demonstrated that it was possible to improve the betulinic acid production in S. cerevisiae while keeping a desirable growth phenotype by optimally balancing the carbon fluxes of the two pathways. Through modulating the expressions of the key genes on betulinic acid and fatty acid pathways, the difference in betulinic acid yield varied largely in the range of 0.01–1.92 mg L？1 OD？1. The metabolic engineering approach used in this study could be extended for synthesizing other triterpenoids in S. cerevisiae.