a State Key Laboratory of Bioactive Substance and Function of Natural Medicines; NHC Key Laboratory of Biosynthesis of Natural Products , Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College , Beijing , P. R. China.
b Biotechnology Core Laboratory , National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health , Bethesda , MD , U.S.A.
Crit Rev Biotechnol. 2019 Mar;39(2):258-271. doi: 10.1080/07388551.2018.1554620. Epub 2019 Jan 2.
Pichia pastoris is extensively used to produce various heterologous proteins. Amounts of biopharmaceutical drugs and industrial enzymes have been successfully produced by fed-batch high-cell-density fermentation (HCDF) of this cell factory. High levels of cell mass in defined media can be easily achieved and therefore large quantities of recombinant proteins with enhanced activities and lower costs can be obtained through HCDF technology. A robust HCDF process makes a successful transition to commercial production. Recently, efforts have been made to increase the heterologous protein production and activity by the HCDF of P. pastoris. However, challenges around selecting a suitable HCDF strategy exist. The high-level expression of a specific protein in P. pastoris is still, at least in part, limited by optimizing the methanol feeding strategy. Here, we review the progress in developments and applications of P. pastoris HCDF strategies for enhanced expression of recombinant proteins. We focus on the methanol induction strategies for efficient fed-batch HCDF in bioreactors, mainly focusing on various stat-induction strategies, co-feeding, and the limited induction strategy. These processes control strategies have opened the door for expressing foreign proteins in P. pastoris and are expected to enhance the production of recombinant proteins.
毕赤酵母被广泛用于生产各种异源蛋白。通过该细胞工厂的分批补料高密度发酵（HCDF），已成功生产出大量的生物制药和工业酶。在限定培养基中可以轻松实现高细胞密度，因此通过 HCDF 技术可以获得具有更高活性和更低成本的大量重组蛋白。稳健的 HCDF 工艺可以成功过渡到商业生产。最近，人们努力通过毕赤酵母的 HCDF 来提高异源蛋白的产量和活性。然而，在选择合适的 HCDF 策略方面仍然存在挑战。在毕赤酵母中高水平表达特定蛋白的能力至少在一定程度上仍然受到优化甲醇补料策略的限制。在这里，我们综述了毕赤酵母 HCDF 策略在增强重组蛋白表达方面的发展和应用的进展。我们重点介绍了生物反应器中高效分批补料 HCDF 的甲醇诱导策略，主要关注各种定态诱导策略、共补料和有限诱导策略。这些过程控制策略为在毕赤酵母中表达外源蛋白开辟了道路，并有望提高重组蛋白的产量。
