The purpose of this chapter is to provide sufficient instruction for the reader to implement fermentation strategies to produce recombinant proteins in the yeastPichia pastoris. P. pastorisutilizes the highly efficient alcohol oxidase 1 gene (AOX1) promoter for high-level expression of foreign genes. TheAOX1promoter is induced by methanol, but repressed in the presence of excess glycerol. Glycerol is readily metabolized byP. pastoris, and high-cell densities can be obtained utilizing this carbon source. The standard fermentation protocol is to growP. pastorisin excess glycerol to repress expression followed by induction of protein production by the addition of methanol after the glycerol is exhausted.
