Evaluation of copper-inducible fungal laccase promoter in foreign gene expression in Pichia pastoris.

The promoter region of copper-inducible laccase gene, LCC1, from Pycnoporus coccineus was explored in the heterologous expression of foreign protein in Pichia pastoris. The promoter region (P) was isolated and used to replace the methanol-inducible AOXI promoter (P) of pPICHOLI-2, an episomal expression vector for P. pastoris, to generate a new copper-inducible expression vector. The promoter activity of P was compared with those of P and P, a copper-inducible promoter of a commercial vector pPICHOLI-C, using a laccase gene as a reporter gene in P. pastoris GS115. Reporter laccase activity of the culture broth reached 182 and 43 units/L for P and P, respectively, after induction with 0.2 mM CuSO at OD = 1 and culture for 120 h at 15°C in complex medium containing 1% glucose. For P activity, yeast cells harboring P-laccase plasmid were cultured for 120 h at 15°C in complex medium with intermittent feeding with 1% methanol every 12 h to avoid methanol toxicity. Laccase activity of culture broth was 124 units/L. Conclusively, P is a new copper-inducible promoter that shows superior performance in terms of efficiency of laccase production compared to commercial vectors. P is additionally superior to P since it does not require laborious feeding with a carbon source. [ABSTRACT FROM AUTHOR]