Co‐Expression of Chaperones for Improvement of Recombinant Geobacillus stearothermophilus Lipase Production in Pichia pastoris
Year: 2025
Type: article
Abstract: Pichia pastoris is among the most popular expression systems for recombinant protein production. The ability to secrete high titers of recombinant proteins is one of its primary advantages. Co-expression of folding-assisting factors is considered one of the strategies to improve protein production with this expression system. In this study, the effect of PDI, KAR2, HAC1, and ERO1 co-expression on Geobacillus stearothermophilus lipase production in P. pastoris was investigated. The chaperones were co-expressed under the regulation of the GAP promoter in clones with different lipase production levels (LPS#2 and LPS#8). Results showed that lipase-PDI co-expression clones have the highest activity. The effect of the other chaperones had varying effects for LPS#2 and LPS#8 clones. LPS#2-PDI and LPS#8-PDI were further analyzed to determine the effect of dimethyl sulphoxide (DMSO). Different concentrations (0.5%, 1%, and 2%) of DMSO were tested. The highest activity was obtained with approximately 1.5-fold activity in the LPS#8-PDI clone with 0.5% and 1% DMSO concentration levels. Comparison of the fermentation kinetic parameters revealed that the activity level of 56.54 U/mL provided with LPS#8 increased to 73.84 U/mL in the LPS#8-PDI clone with PDI co-expression, and when PDI co-expression was combined with DMSO conditions (0.5%), it reached 114.46 U/mL. It was observed that the productivity (U/g wet cell/g substrate/h) of the LPS#8 clone was increased 1.27-fold with LPS#8-PDI and 2.05-fold with LPS#8-PDI-DMSO conditions. In future studies, it is possible to achieve higher protein production levels by optimization at the fermenter scale.
DOI: 10.1002/bab.70017
Author: Fidan Erden‐Karaoglan
Institution: Erzincan Binali Yıldırım University
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: bronze
OpenAlex ID: W4411908978