Nyberg G B, Balcarcel R R, Follstad B D, Stephanopoulos G, Wang D I
Department of Chemical Engineering, Biotechnology Process Engineering Center, Room 56-469C, Massachusetts Institute of Technology, MIT, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
Biotechnol Bioeng. 1999 Feb 5;62(3):324-35.
Metabolic flux analysis is a useful tool for unraveling relationships between metabolism and cell function. Material balancing can be used to provide estimates of major metabolic pathway fluxes, provided all significant metabolite uptake and production rates are measured. Potential sources of metabolizable material in many serum-free media formulations are low molecular weight digests of biological material such as yeast extracts and plant or animal tissue hydrolysates. These digests typically contain large amounts of peptides, which may be utilized as amino acids. This article demonstrates the need for accounting for amino acids liberated from peptides in order to accurately estimate pathway fluxes in Chinese hamster ovary cells grown in a complex (hydrolysate containing) medium. A simplified model of central carbon metabolism provides the framework for analyzing external metabolite measurements. Redundant measurements are included to ensure the consistency of data and assumed biochemistry by comparing redundant measurements with their predicted values from a minimum data set, and by expressing the degree of agreement using a statistical "consistency index." The consistency index tests whether redundancies are satisfied within expected experimental error. For chemostat steady states of CHO cultures grown in a hydrolysate-supplemented medium, consistent data were obtained only when amino acids liberated from peptides were taken into account.
代谢通量分析是一种用于揭示代谢与细胞功能之间关系的有用工具。如果能够测量所有重要代谢物的摄取和产生速率，那么物料平衡可用于估算主要代谢途径的通量。在许多无血清培养基配方中，可代谢物质的潜在来源是生物材料的低分子量消化产物，如酵母提取物以及植物或动物组织水解产物。这些消化产物通常含有大量的肽，这些肽可作为氨基酸被利用。本文证明，为了准确估算在复杂（含水解产物）培养基中生长的中国仓鼠卵巢细胞的途径通量，需要考虑从肽中释放出的氨基酸。一个简化的中心碳代谢模型为分析外部代谢物测量提供了框架。通过将冗余测量值与其从最小数据集预测的值进行比较，并使用统计“一致性指数”来表示一致程度，从而纳入冗余测量以确保数据的一致性和假设的生物化学过程。一致性指数检验在预期实验误差范围内冗余是否得到满足。对于在添加了水解产物的培养基中生长的CHO培养物的恒化器稳态，只有在考虑从肽中释放出的氨基酸时才能获得一致的数据。
