[Effect of signal peptides on the expression of laccase in Pichia pastoris]

Abstract

Objective: Expression level of laccase POXA1c would be increased by screening effective signal peptide in Pichia pastoris.

Methods: According to 2D-gel and profile of P. pastoris genome sequence, seven signal peptides from high secreted endogenous proteins of P. pastoris X-33 were chosen to evaluate their secreted ability by using POXA1c as reporter protein.

Results: Compared with POXA1c's native signal peptide, the signal peptide of repressible acid phosphatases PHO5 and lectin-like protein FLO10 showed 2. 5-fold and 2-fold increase of laccase activity. Furthermore, PHO5-αpro and FLO10-αpro were constructed by fusing signal peptide of PHO5 and FLO10 with pro-peptide of α-MF respectively. The laccase activity under the leading of FLO10-αpro and PHO5-αpro showed 3-fold and 3. 5- fold laccase activity higher than native signal peptide, and showed 20% and 40% increase compared with saccharomyces cerevisiae α-MF signal respectively.

Conclusion: Signal peptides from high secreted endogenous proteins of P. pastoris X-33 could be effectively used to lead laccase expression in P. pastoris. The activity of POXA1c under the leading of the PHO5-αpro signal peptide was 57.98 U/mL after high density fermentation.