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 methanol-complex medium. Co-expression of the HAC1ⁱ gene enhanced the expression level of the heterogeneously N-glycosylated 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.

在这项研究中，评估了通过共表达HAC 1 ⁱ基因激活未折叠蛋白反应（UPR）对重组巴斯德毕赤酵母GS115中铜绿假单胞菌弹性蛋白酶（rPAE）产生的影响。结果表明，HAC 1 ⁱ基因的表达显着提高了巴斯德毕赤酵母GS115中Kar2p（UPR激活的标志）的水平，表明UPR的激活。该基因不影响酵母在甘油复合缓冲液中的生长，但刺激了酵母在甲醇复合缓冲液中的生长。HAC 1 ^i的共表达该基因增强了rPAE的异种N-糖基化形式的表达水平，因为巴斯德毕赤酵母GS115 / HAC1中表达的rPAE的各种糖型上清液中的酪蛋白水解活性比对照菌株增加了1.8-3.9倍巴斯德毕赤酵母GS115。每24小时添加0.5、1.0和2.0％甲醇时，可观察到该基因共表达对rPAE产生的刺激作用，因为表达野生型rPAE的巴斯德毕赤酵母GS115 / HAC1上清液的酪蛋白水解活性为rPAE。在相应的甲醇浓度下增加了3.3倍，1.9倍和1.7倍。此外，通过HAC 1 ^i的共表达激活UPR基因表达增强了20、24和28°C下巴斯德毕赤酵母中rPAE的分泌，这是因为表达野生型rPAE的巴斯德毕赤酵母GS115 / HAC1上清液的酪蛋白分解活性是其的2.3、2.1和2.8倍。测试温度。