Jin S, Ye K, Shimizu K
Department of Biochemical Engineering and Science, Kyushu Institute of Technology, Fukuoka, Japan.
J Biotechnol. 1997 May 9;54(3):161-74. doi: 10.1016/s0168-1656(97)01694-5.
A stoichiometric flux balancing analysis was applied to the recombinant yeast cultivation to examine the cellular physiology and relationship between the production of heterologous protein and metabolic fluxes. The fluxes in the metabolic pathway within a recombinant S. cerevisiae grown on galactose alone or mixtures of galactose and ethanol medium were calculated. It is found that an amplification of the PP (Pentose Phosphate) pathway activity resulted in an improvement of the foreign protein expression and cell yield on ATP. The carbon source used for biosynthesis from TCA cycle in the exponential growth phase was 2 and 5-fold higher, respectively, as compared with that in the late exponential growth phase and stationary phase in batch culture with galactose minimum medium. The metabolism of ethanol together with galactose in the recombinant cell looks like increasing the flux from Acetyl-CoA to TCA cycle, and amplifying the flux directing the synthesis of various kinds of precursors such as amino acids and nucleic acid which are necessary for production of a foreign protein. Metabolic flux distribution analysis also shows that the ATP synthesis rate under substrate-level phosphorylation in the mixed carbon source cultivation was lower than that in the sole carbon source (galactose) during the expression of foreign protein. However, the total ATP production rate was higher in the mixed carbon source cultivation.
应用化学计量通量平衡分析方法对重组酵母培养进行研究，以考察细胞生理学以及异源蛋白生产与代谢通量之间的关系。计算了在仅以半乳糖或半乳糖与乙醇混合培养基上生长的重组酿酒酵母代谢途径中的通量。研究发现，磷酸戊糖（PP）途径活性的增强导致外源蛋白表达和基于ATP的细胞产量提高。在指数生长期，与在含半乳糖基本培养基的分批培养中的指数生长后期和稳定期相比，三羧酸循环用于生物合成的碳源分别高出2倍和5倍。重组细胞中乙醇与半乳糖的共同代谢似乎增加了从乙酰辅酶A到三羧酸循环的通量，并放大了导向合成外源蛋白生产所需的各种前体（如氨基酸和核酸）的通量。代谢通量分布分析还表明，在表达外源蛋白期间，混合碳源培养中底物水平磷酸化下的ATP合成速率低于单一碳源（半乳糖）培养中的速率。然而，混合碳源培养中的总ATP产生速率更高。
