文献类型：期刊文献

中文题名：毕赤酵母表达全长单克隆抗体的设计及优化

英文题名：Expression design and optimization of full-length monoclonal antibodies in Pichia pastoris

作者：叶凯雄[1];龚秀龙[1];徐名强[1];钱芷兰[1];刘启[1];蔡孟浩[1]

机构：[1]华东理工大学生物工程学院生物反应器国家重点实验室,上海200237

年份：2025

卷号：38

期号：4

起止页码：427

中文期刊名：中国生物制品学杂志

外文期刊名：Chinese Journal of Biologicals

基金：国家自然科学基金青年基金(32201206);中国博士后科学基金面上资助(2022M711146)。

语种：中文

中文关键词：单克隆抗体;毕赤酵母;分子伴侣

外文关键词：Monoclonal antibodies(mAbs);Pichia pastoris;Molecular chaperone

摘要：目的利用毕赤酵母底盘宿主表达单克隆抗体,并探索多种表达优化策略以提高抗体产量。方法以毕赤酵母为底盘细胞,用甲醇诱导型系统PAOX1和乙醇诱导型系统E1分别尝试多种单克隆抗体蛋白的重组表达,并通过分子伴侣过表达和培养基组分优化进一步提升产量后,经3 L反应器高密度发酵培养验证重组菌株生产能力。结果乙醇诱导表达的依普奈珠单克隆抗体产量可达12.4 mg/L。分子伴侣Ppi1和Biph的过表达分别使依普奈珠单克隆抗体产量提升约22.5%和23.0%;添加1.0 g/L硫酸铵、0.05 mol/L磷酸钾缓冲对和80 mmol/L苏氨酸也分别使依普奈珠单克隆抗体产量提升约57.1%、25.8%和58.1%。在3 L生物反应器水平,Biph过表达菌株的完整抗体产量相较于摇瓶水平提升约5.9倍,最高产量达74.1 mg/L。结论依普奈珠单克隆抗体在毕赤酵母底盘宿主中可成功表达,并且通过多种表达调控策略能够有效提升抗体产量,为毕赤酵母表达单克隆抗体提供了新的思路和方向。
Objective To express monoclonal antibodies(mAbs)in Pichia pastoris chassis host and explore various expression optimization strategies to improve the yield of antibodies.Methods Pichia pastoriswas utilized as the chassis cell for the recombinant expression of various mAbs using both methanol-induced system PAOX1 and ethanol-induced system E1.Molecular chaperone overexpression and media component optimization were employed to further enhance the production of mAbs.The productivity of the recombinant strains was then evaluated at a 3 L reactor scale.Results Ethanol-induced expression of Eptinezumab resulted in a yield of 12.4 mg/L.Overexpression of molecular chaperones Ppi1 and Biph increased Eptinezumab production by approximately 22.5%and 23.0%,respectively.Additionally,the supplementation of1.0 g/L ammonium sulfate,the adjustment of 0.05 mol/L potassium phosphate buffer,and the addition of 80 mmol/L threonine individually enhanced Eptinezumab yields by approximately 57.1%,25.8%,and 58.1%,respectively.At the 3 L bioreactor scale,the Biph-overexpressing strain exhibited a remarkable increase in complete antibody production,achieving a maximum yield of 74.1 mg/L,which was approximately 5.9 times higher than that obtained at the flask level.Conclusion Eptinezumab can be successfully expressed in the Pichia pastoris chassis host,and the application of various expression optimization strategies has effectively enhanced the production of the antibody.This study provides novel insights and directions for the expression of monoclonal antibodies in Pichia pastoris.