Combinatorial library strategy for strong overexpression of the lipase from Geobacillus thermocatenulatus on the cell surface of yeast Pichia pastoris

Abstract

The yeast cell surface display technique allows for the expression of a target protein on the yeast cell surface and has many applications such as the immobilization of enzymes and the development of biosensors. To increase the expression of the BTL2, a lipase from Geobacillus thermocatenulatus, on the cell surface of yeast Pichia pastoris, we developed a combinatorial library strategy for selecting appropriate expression cassette comprising sequences encoding a promoter, secretion signal, mature BTL2, anchoring protein, and terminator. The transformant GS115/D90, which comprised P. pastoris ENO1 promoter sequence, Hansenula polymorpha GAS1 secretion signal sequence, and Saccharomyces cerevisiae GAS1 anchoring protein gene, exhibited 5-fold higher lipase activity compared to the control strain harboring a conventional expression cassette. Using the developed strategy, an appropriate expression cassette for the strong overexpression of target proteins on the cell surface of yeast could be rapidly and easily obtained.