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

•Enzymes can be displayed on yeast cell surface by cell surface display technique.•Combinatorial library strategy for the lipase overexpression in yeast was developed.•GS115/D90 exhibited 5-fold higher lipase activity was obtained by the strategy.•GS115/D90 comprised ENO1 promoter, GAS1 secretion signal, and GAS1 anchoring protein.•Overexpression on yeast cell surface could be achieved easily by the strategy.

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.

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