Purification of MBP‐tagged proteins using new prepacked columns

Abstract

The success of protein production and subsequent analysis are dependent on the expression levels, solubility and purification of the protein. Maltose Binding Protein (MBP) is often used as an affinity fusion tag. Several studies have shown that MBP‐tagged proteins have high solubility compared to other tagged proteins.The new MBPTrap™ HP consists of an affinity medium, Dextrin Sepharose™ High Performance, packed in 1‐ml and 5‐ml HiTrap™ columns. The medium has high specificity and capacity. Additionally, it can withstand harsh Cleaning In Place (CIP) procedures, like sodium hydroxide, without losing binding capacity. The HiTrap format enables automated protocols to be used on ÄKTAdesign™ systems as well as with syringe or laboratory pump.In this work, MBP‐tagged proteins from E. coli were purified on prepacked 1‐ml or 5‐ml MBPTrap HP columns. The purity of the eluted target proteins was greater than 90 %. A stability study was performed, six purification runs each followed by a CIP run (0.5 M NaOH) were made on the same column. The results show no significant change in yield or purity. The results of an automated two step purification on ÄKTAxpress™ is shown. The affinity chromatography step on MBPTrap HP was followed by a gel filtration step.In summary, the results show that the prepacked MBPTrap HP column has high binding capacity and the eluted target protein is very pure. Furthermore, the medium is stable in 0.5 M NaOH.