Proteomic Characterisation of a Glucose-Limited CHO Perfusion Process–Analysis of Metabolic Changes and Increase in Productivity

Abstract

In order to identify proteins that are engaged in the increase of cell specific productivity and the associated metabolic shift under glucose-limiting conditions, we examined three different phases of a perfusion cultivation process of CHO-MUC1-IgG2a cells by means of proteomic analysis using 2D-DIGE-technology followed by MS-based protein identification.

Cell extracts of exemplary time points for the three different fermentation stages were labelled with CyDye DIGE Fluor minimal dyes (GE Healthcare) and separated on 2D-PAGE in four replicates, including a duplicate dye swap.

Up to now, a total of 1064 analysed 2D-spots using Delta 2D software (Decodon, Greifswald) revealed about 600 proteins that demonstrate a more than twofold change in expression levels throughout the fermentation stages with a significance of p < 0.02 according to students t-test.

Identified proteins belong, among others, to functional protein groups like energy metabolism, protein biosynthesis, stress/chaperones, cytoskeleton and cell signalling.