Purification of untagged retroviral integrases by immobilized metal ion affinity chromatography

We have developed a simple protocol for the purification of untagged retroviral integrases expressed in bacterial cells. The method takes advantage of the inherent ability of the proteins to bind metal ions. The protocol involves an initial enrichment of the protein in the pellet fraction following centrifugation of the lysate after cell lysis. Integrase is then solubilized from the pellet at high salt conditions (1 M) with detergent and applied to a nickel-charged iminodiacetic acid-Sepharose column. The enzyme is eluted from the column with imidazole. The resulting protein, which is 70-80% homogeneous, is subsequently purified to homogeneity on a heparin-Sepharose column. The two-column protocol is easily completed in a day and yields approximately 2 mg of enzymatically active protein per gram of wet cell paste.