Interloping frontiers of systems biology : mass spectrometry in bioprocesses optimization [article]

Andrés Sánchez-Kopper, Universität Stuttgart, Universität Stuttgart
2017
System biology as the understanding and prediction of how intracellular machinery works, needs new technological support in order to observe cell metabolism at all its levels. From organism genome to its transcription to proteins and how these proteins regulate metabolite pools as the final response to determined stimuli, are needed to have a close look into specific metabolic states. Mass spectrometry is used here to obtain intracellular information at a metabolomics, peptidomics and
more » ... omics and proteomics level for specific questions concerning bioprocesses optimization. First, a ZIC-HILIC tandem mass spectrometric method was established to allow the intracellular quantitation of about 50 polar metabolites of the central carbon metabolism without extra derivatization steps, increasing detection limits using alkaline mobile phase, considering "wrong-way-around" ionization, as a basis to perform metabolic flux analysis, needed to evaluate metabolic engineering approaches over titer production. Also, culture performance boosting dipeptide uptake was for the first time revealed by intracellular dipeptide pools quantification in CHO cells, by means of high-resolution mass spectrometry, with which tracking its uptake rates, shows the presence of two possible mechanisms for dipeptide uptake in CHO-DP12 cells. Dipeptide metabolization inside the cell was additionally revealed, as amino acids coming from the dipeptides uptake are directly metabolized or expelled out of the cell proving that, in order to optimize mammalian cells bioprocesses, by means of dipeptide medium supplementation, there is a need to understand specific dipeptide uptake and metabolization. Furthermore, protein turnover of a reference molecular antibody, produced in CHO cells, was also chased by a novel methodology. Out of the traditional stable isotope labeling with amino acids (SILCA), production-like conditions were evaluated, the intracellular mAb degradation was for the first time partially quantified, as intracellular peptides produced by Anti-interleukin [...]
doi:10.18419/opus-9094 fatcat:fgsdpu3onjduraafcoieexksu4