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Abstract
Expression of recombinant proteins using affinity tags is a commonly used purification strategy. Histidine and glutathione‐S transferase (GST) are the most common and versatile of these tags. Histidine tags are small and therefore less disruptive to the protein structures and functions. GST tags enhance solubility of the target protein, hence improving the yield.Manual purification of tagged proteins often face issues with time to purify, low reproducibility and low yield etc. In addition a researcher who is purifying manually needs to spend time in setting‐up the run, sample loading, collection and measuring the final protein concentration.We describe here the purification Histidine‐tagged and GST‐tagged protein using ÄKTA start with Frac30 fraction collector and UNICORNTM start system control software.We purified an N‐terminal Histidine tagged protein expressed in E.coli on a pre‐packed HisTrapTM FF 1mL column. The purification method was completely automated for unattended operation by applying large volume of sample using pump and collecting peak fractions using Frac30 fraction collector. We have consistently obtained high levels of purity (> 95%) of the tagged protein, which is 1.6 fold higher when compared to a conventional manual purification method.In the second example, we describe the purification of GST tagged protein expressed in E.coli on a pre‐packed GSTrapTM FF 1ml column using ÄKTA start with Frac30 fraction collector. The target protein was purified using pre‐defined method workflow available from the instrument display. The result file that was stored on USB memory stick is imported for viewing and report generation using UNICORN start software.ÄKTATM start is the newly developed laboratory scale protein purification system that can address the challenges faced during manual purification. Automated purification of tagged proteins has showed higher yields and purity with high consistency.
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