Similarity of the Escherichia coli proteome upon completion of different biopharmaceutical fermentation processes.
A comprehensive view of the physiological state of Escherichia coli cells at the completion of fermentation processes for biopharmaceutical production was attained via two-dimensional gel electrophoretic analysis of cellular proteins. For high cell density fermentations in which phosphate is depleted to induce recombinant protein expression from the alkaline phosphatase promoter, proteome analysis confirms that phosphate limitation occurs. Known phosphate starvation inducible proteins are observed at high levels; these include the periplasmic phosphate binding protein and the periplasmic phosphonate binding protein. The phn (EcoK) locus of these E. coli K-12 strains remains cryptic, as demonstrated by failure to grow with phosphonate as the sole phosphorus source. Proteome analysis also provided evidence that cells utilize alternative carbon and energy sources during these fermentation processes. To address regulatory issues in the biopharmaceutical industry, comparative electrophoretic analyses were conducted on a qualitative basis for four different fermentation processes. Using this approach, the protein profiles for these processes were found to be highly similar, with the vast majority (85-90%) of proteins detected in all profiles. The observed similarity in proteomes suggests that multiproduct host cell protein immunoassays are a feasible means of quantifying host-derived polypeptides from a variety of biopharmaceutical fermentation processes.
Champion
K M
KM
Department of Analytical Chemistry, Genentech, South San Francisco, CA 94080, USA. kmc@gene.com
Nishihara
J C
JC
Joly
J C
JC
Arnott
D
D
D001702:Biopharmaceutics; D018821:CD18 Antigens / Q000378:metabolism; D002244:Carbon / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D016228:Endothelial Growth Factors / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D005285:Fermentation; D006360:Heat-Shock Proteins / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D008222:Lymphokines / Q000378:metabolism; D063065:Organophosphonates / Q000378:metabolism; D020543:Proteome / Q000737:chemistry* / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D018821:CD18 Antigens; D016228:Endothelial Growth Factors; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D008222:Lymphokines; D063065:Organophosphonates; D020543:Proteome; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D007334:Insulin-Like Growth Factor I; D002244:Carbon