<jats:title>ABSTRACT</jats:title> <jats:p> ( <jats:italic>E</jats:italic> , <jats:italic>E</jats:italic> , <jats:italic>E</jats:italic> )-Geranylgeraniol (GGOH) is a valuable starting material for perfumes and pharmaceutical products. In the yeast <jats:italic>Saccharomyces cerevisiae</jats:italic> , GGOH is synthesized from the end products of the mevalonate pathway through the sequential reactions of farnesyl diphosphate synthetase (encoded by the <jats:italic>ERG20</jats:italic> gene), geranylgeranyl diphosphate synthase (the <jats:italic>BTS1</jats:italic> gene), and some endogenous phosphatases. We demonstrated that overexpression of the diacylglycerol diphosphate phosphatase ( <jats:italic>DPP1</jats:italic> ) gene could promote GGOH production. We also found that overexpression of a <jats:italic>BTS1</jats:italic> - <jats:italic>DPP1</jats:italic> fusion gene was more efficient for producing GGOH than coexpression of these genes separately. Overexpression of the hydroxymethylglutaryl-coenzyme A reductase ( <jats:italic>HMG1</jats:italic> ) gene, which encodes the major rate-limiting enzyme of the mevalonate pathway, resulted in overproduction of squalene (191.9 mg liter <jats:sup>−1</jats:sup> ) rather than GGOH (0.2 mg liter <jats:sup>−1</jats:sup> ) in test tube cultures. Coexpression of the <jats:italic>BTS1</jats:italic> - <jats:italic>DPP1</jats:italic> fusion gene along with the <jats:italic>HMG1</jats:italic> gene partially redirected the metabolic flux from squalene to GGOH. Additional expression of a <jats:italic>BTS1</jats:italic> - <jats:italic>ERG20</jats:italic> fusion gene resulted in an almost complete shift of the flux to GGOH production (228.8 mg liter <jats:sup>−1</jats:sup> GGOH and 6.5 mg liter <jats:sup>−1</jats:sup> squalene). Finally, we constructed a diploid prototrophic strain coexpressing the <jats:italic>HMG1</jats:italic> , <jats:italic>BTS1</jats:italic> - <jats:italic>DPP1</jats:italic> , and <jats:italic>BTS1</jats:italic> - <jats:italic>ERG20</jats:italic> genes from multicopy integration vectors. This strain attained 3.31 g liter <jats:sup>−1</jats:sup> GGOH production in a 10-liter jar fermentor with gradual feeding of a mixed glucose and ethanol solution. The use of bifunctional fusion genes such as the <jats:italic>BTS1</jats:italic> - <jats:italic>DPP1</jats:italic> and <jats:italic>ERG20</jats:italic> - <jats:italic>BTS1</jats:italic> genes that code sequential enzymes in the metabolic pathway was an effective method for metabolic engineering.