Monday, August 13, 2012
Columbia Hall, Terrace Level (Washington Hilton)
Mathilde Mosser
1, Romain Kapel
1, Laurent-Michel Bonanno
2, Jim Alferman
3,
Alain Sourabié2, Eric Olmos
1, Isabelle Chevalot
1, Ivan Marc
1 and Annie Marc
1, (1)Laboratoire Réactions et Génie des Procédés, Université de Lorraine, Vandoeuvre-lès-Nancy, France, (2)Biotechnology Application Lab, Bio Springer, Maisons-Alfort, France, (3)Bio Springer North America, Montreal, QC, Canada
Protein hydrolysates from yeasts or plants are currently used in many microbial systems, e.g. biopharmaceutical productions, as supplements in order to reduce or eliminate serum-based media requirements. Yeast extract (YE), the water soluble fraction of molecules yielded after full autolysis, is a complex mixture mainly composed of free amino acids, peptides, carbohydrates, nucleic acids, vitamins and minerals. It was shown to improve cell growth and viability as well as recombinant protein productivity. The aim of this study was to i) identify and ii) characterize the physicochemical properties of YE derived molecules, enhancing CHO cell growth and protein yield, by using chromatographic fractionation.
A nanofiltrated YE (nYE) was first fractionated by preparative chromatography, either with anion exchange (AEC), hydrophobic interaction (HIC) or size exclusion (SEC). Each fraction was then added in a control chemically defined medium to assess its impact on CHO cell growth.
Results mainly highlighted that AEC was the most selective separation process to purify nYE in one step without decreasing cell growth. A three-step chromatographic process including successive AEC, HIC, and SEC was further developed to refine the physicochemical properties of nYE compounds. Among fractions that triggered similar cell growth promoting properties compared to nYE, one also improved IgG specific production. It was composed of cationic and hydrophilic peptides with a great proportion of lysine and arginine, low quantities of polysaccharides and no nucleic acids.
In this study, we demonstrate that a specific mixture of nYE molecules is most probably involved in YE growth promoting activity on CHO.
