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KMID : 1094720200250020302
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Biotechnology and Bioprocess Engineering
2020 Volume.25 No. 2 p.302 ~ p.307
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Activation of the Unfolded Protein Response via Co-expression of the HAC1i Gene Enhances Expression of Recombinant Elastase in Pichia pastoris
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Han Minghai
Wang Weixian
Zhou Jianli
Gong Xun
Xu Cunbin
Li Yinfeng
Li Qiang
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Abstract
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The effects of activation of the unfolded protein response (UPR) via co-expression of the HAC1i gene on the production of the recombinant Pseudomonas aeruginosa elastase (rPAE) in Pichia pastoris GS115 were evaluated in this study. The results showed that expression of the HAC1i gene significantly increased the level of Kar2p (a hallmark of UPR activation) in P. pastoris GS115, demonstrating activation of the UPR. This gene did not affect the growth of yeast in the buffered glycerol-complex medium but stimulated its growth in the buffered methanolcomplex medium. Co-expression of the HAC1i gene enhanced the expression level of the heterogeneously Nglycosylated forms of rPAE, as the caseinolytic activity in the supernatant of the various glycoforms of rPAE expressed in P. pastoris GS115/HAC1 was increased 1.8?3.9-fold compared to that in the control strain P. pastoris GS115, respectively. The stimulating effects of co-expression of the gene on rPAE production were observed when 0.5, 1.0, and 2.0% methanol were added every 24 h, as the caseinolytic activity of supernatants of P. pastoris GS115/HAC1 expressing wild-type of rPAE was increased 3.3-, 1.9-, and 1.7-fold at the corresponding methanol concentration. Further, activation of UPR via co-expression of the HAC1i gene enhanced rPAE secretion in P. pastoris at 20, 24, and 28°C, as the caseinolytic activity of supernatants of P. pastoris GS115/HAC1 expressing wild-type rPAE was increased 2.3-, 2.1-, and 2.8-fold over the tested temperatures.
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KEYWORD
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unfolded protein response, Pichia pastoris, protein expression, HAC1
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