Purification of an elastin-like fusion protein by microfiltration

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Abstract
This article describes a simple and potentially scalable microfiltration method for purification of recombinant proteins. This method is based on the fact that when an elastin‐like polypeptide (ELP) is fused to a target protein, the inverse phase transition behavior of the ELP tag is imparted to the fusion protein. Triggering the phase transition of a solution of the ELP fusion protein by an increase in temperature, or isothermally by an increase in salt concentration, results in the formation of micron‐sized aggregates of the ELP fusion protein. In this article, it is shown that these aggregates are efficiently retained by a microfiltration membrane, while contaminating E. coli proteins passed through the membrane upon washing. Upon reversing the phase transition by flow of Milli‐Q water, soluble, pure, and functionally active protein is eluted from the membrane. Proof‐of principle of this approach was demonstrated by purifying a fusion of thioredoxin with ELP (Trx‐ELP) with greater than 95% recovery of protein and with greater than 95% purity (as estimated from SDS–PAGE gels). The simplicity of this method is demonstrated for laboratory scale purification by purifying Trx‐ELP from cell lysate using a syringe and a disposable microfiltration cartridge. The potential scalability of this purification as an automated, continuous industrial‐scale process is also demonstrated using a continuous stirred cell equipped with a microfiltration membrane.

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