High-level expression of the Penicillium notatum glucose oxidase gene in Pichia pastoris using codon optimization
High-level expression of the Penicillium notatum glucose oxidase gene in Pichia pastoris using codon optimization
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DOI：
10.1007/s10529-011-0790-6
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发表时间：
2012-03-01
影响因子：
2.7
通讯作者：
Zhang, Wei
中科院分区：
文献类型：
--
作者：
Gao, Zhaowei;Li, Zhuofu;Zhang, Wei
利用巴斯德毕赤酵母表达了诺氏青霉葡萄糖氧化酶(GOD)基因。该基因全长1,815个碱基，编码604个氨基酸。重组GOD-W在35-40℃、pH 6.2条件下具有最高活性，在pH 3-7范围内稳定，在50℃孵育1h后，GOD-W的活性保持在最大值75%。为了实现重组GOD在巴斯德毕赤酵母中的高效表达，共突变了272个核苷酸，涉及228个残基，与巴斯德毕赤酵母的密码子偏好性一致。优化后的重组GOD-M在3号L发酵罐中的发酵产量为615U ml(-1)(2.5g L蛋白)，比GOD-W(148U ml(-1))高出410%，是面包烘焙行业的低成本替代品。
The glucose oxidase (GOD) gene from Penicillium notatum was expressed in Pichia pastoris. The 1,815 bp gene, god-w, encodes 604 amino acids. Recombinant GOD-w had optimal activity at 35-40 degrees C and pH 6.2 and was stable, from pH 3 to 7 maintaining >75% maximum activity after incubation at 50 degrees C for 1 h. GOD-w worked as well as commercial GODs to improve bread making. To achieve high-level expression of recombinant GOD in P. pastoris, 272 nucleotides involving 228 residues were mutated, consistent with the codon bias of P. pastoris. The optimized recombinant GOD-m yielded 615 U ml(-1) (2.5 g protein l(-1)) in a 3 l fermentor-410% higher than GOD-w (148 U ml(-1)), and thus is a low-cost alternative for the bread baking industry.