Abstract:

As green and sustainable low-carbon feedstocks without competition with people for food, methanol and other C₁ compounds have great potential for advancement of biomanufacturing. Pichia pastoris (P. pastoris),as an industrial strain capable of naturally utilizing methanol, has a long history of research and has been widely employed in the biomanufacturing of various recombinant proteins. It has become an excellent chassis cell factory for methanol utilization. In recent years, there has been progress in understanding the microbial physiology, methanol metabolism networks, protein secretion expression pathways, and the identification of key components in P. pastoris. However, several critical challenges remain that limit its efficient application, such as limited understanding of the complex mechanisms of cell toxicity caused by methanol and its metabolites, insufficient capacity to explore “biological dark matters” related to cell robustness and methanol utilization, and restricted engineering approaches for efficiently converting methanol into target proteins. This review summarizes the research progress on the bioconversion of methanol into recombinant proteins by P. pastoris, with the emphasis on discussing how to optimize the methanol conversion and protein expression in P. pastoris cell factory. Finally, we address the challenges encountered in the recombinant protein production from methanol in P. pastoris and provide an outlook on its future biotechnology innovation and potential application.

Key words: methanol, Pichia pastoris, recombinant proteins, metabolic engineering, synthetic biology

摘要：

甲醇等一碳化合物作为生物制造的一种绿色可持续低碳原料，不存在与人争粮的问题，具有巨大的应用发展潜力。能够天然利用甲醇的毕赤酵母研究历史长，已被广泛应用于多种重组蛋白的生物制造中，成为甲醇利用性能优良的底盘细胞工厂。近年虽然对毕赤酵母的生理学、甲醇代谢网络、蛋白分泌表达途径及关键元件挖掘开展了大量的研究，积累了大量知识，但仍存在影响其高效应用的一系列关键问题，如甲醇及其代谢物对细胞毒性的复杂影响机制认知有限、细胞鲁棒性及甲醇利用相关的“生命暗物质”挖掘能力不足、甲醇高效转化成目标蛋白的工程化手段有限等。本文综述了国内外有关毕赤酵母生物转化甲醇合成重组蛋白的研究现状，重点总结讨论了毕赤酵母细胞工厂转化甲醇及蛋白表达优化策略，最后探讨了当前毕赤酵母生物转化甲醇生产重组蛋白过程中面临的挑战，并对其未来的生物技术创新和应用进行了展望。

关键词: 甲醇, 毕赤酵母, 重组蛋白, 代谢工程, 合成生物学