Quantitative Analysis of Proteome Dynamics in Chinese Hamster Ovary cells

The overall goal of this research was to better understand the mechanisms underlying the physiology of CHO cells, the most important mammalian host for recombinant protein production. The publication of complete genome of CHO cells allowed the use of mass-spectrometry based proteomic tools to study protein expression. Among several different sample preparation methods for mass spectrometry, in-gel trypsin digest and FASP were found to be the most robust and optimal for high-coverage CHO proteome analysis. Global changes in protein expression between exponential and stationary phases were determined using SILAC for parental GS K-O and producing E22 cell lines. >4000 proteins have been quantified and more than 100 proteins have been statistically differentiated. Proteins up-regulated in exponential phase control cell cycle and DNA replication, while proteins up-regulated in the stationary phase are involved in stress response and signalling, making them interesting targets for cellular engineering. In addition to quantifying relative changes in protein expression between two phases of cell culture, more than 4000 protein copy numbers were calculated for parental and producing cell lines using TPA method. Protein turnover, described as the balance between protein synthesis and degradation, was calculated for >3000 cellular proteins. Combining these two parameters together allowed determination of top 10 proteins corresponding to 20% of global turnover rate. Production of monoclonal antibody was top priority, causing metabolic burden on cells. KEGG and GO annotation suggests that 600 up-regulated proteins in E22 producing cell line explained their clonal selection based on highest growth and productivity. Interestingly, there was no major differences found between amino acid and codon usage between parental and producing cell lines. In summary, a large-scale proteomic data set containing qualitative, quantitative and dynamic information on protein expression for industrially relevant CHO cell lines.