Effect of Temperature on the Secretory Expression of Porcine Insulin Precursor and Physiological Characterization of Multicopy P. pastoris

The impact of cultivation temperature on the physiology and productivity of porcine insulin precursor (PIP) protein secreting P. pastoris strains was investigated. The multicopy PIP producing P. pastoris strains A2 and A3 were cultivated at 25 ℃ and 30 ℃. Compared with those at 30 ℃， PIP production of A2 and A3 at 25 ℃ was increased by 40% and 32%, respectively. The viability of recombinant A3 cells was increased by 20%, and the transcriptional levels of KAR2, PDI1, GLR, and TRR1 in A3 cells were reduced by 16%-35% compared with those cultivated at 30 ℃. Thus, protein folding stress and oxidative stress responses in the highcopy strain cells are generally decreased at 25 ℃, leading to a 40% increase in heterologous protein production by multicopy recombinart P. pastoris host cells.