Interfacing Pichia pastoris cultivation with expanded bed adsorption
© Jahic et al; licensee BioMed Central Ltd. 2006
Published: 10 October 2006
For improved interfacing of the Pichia pastoris fed-batch cultivation process with expanded bed adsorption (EBA) technique, a modified cultivation technique was developed. The modification included the reduction of the medium salt concentration, which was then kept constant by regulating the medium conductivity at low value (about 8 mS cm-1) by salt feeding. Before loading, the low conductivity culture broth was diluted only to reduce viscosity, caused by high cell density. The concept was applied to a one-step recovery and purification procedure for a fusion protein composed of a cellulose binding module (CBM) from Neocallimastix patriciarum cellulase 6A fused to lipase B from Candida antarctica (CALB).
Our achievements in the modified cultivation stage that increased the quality of the feedstock for the separation stages were: higher target protein to total protein concentration ratio in the culture broth, low protease activities, and lower salt concentration and conductivity. The low salt and conductivity should also make the feedstock more suitable for loading on ion exchanger EBA media. These achievements made the expanded bed adsorption technique more efficient for initial recovery of CBM-CALB in particular but should also make the EBA technique more attractive for the recovery of other recombinant proteins from P. pastoris system.
This work is part of the BiMac Enzyme Factory programme and financed by the Södra Skogsägarnas Stiftelse for Forskning, Utveckling och Utbildning.
This article is published under license to BioMed Central Ltd.