Abstract
A purification system was constructed with the N-segment of the Npu DnaE split intein as an affinity ligand immobilized onto an epoxy-activated medium and the C-segment used as the cleavable tag fusing target protein. The affinity properties of C-tagged proteins adsorbed on IN affinity chromatography medium were studied with GFP as a model target protein. The saturated adsorption capacity and dynamic adsorption capacity reached 51.9-21.0mgmL-1, respectively. With this system, two model proteins, GFP and alcohol dehydrogenase (ADH), has been successfully taglessly purified with regulation of Zn2+ and DTT. The yield, purification factor and purity of purified tagless GFP reached 39, 11.7 and 97%, respectively; while these values for purified tagless ADH were 38.2, 6.8 and 91%, respectively. These results showed that the system for Npu DnaE split intein-mediated affinity adsorption and in situ cleavage is a potential platform for recombinant protein production.
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