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  • Title: Separation of hexahistidine fusion proteins with immobilized metal ion affinity chromatographic (IMAC) sorbents derived from M(N+)-tacn and its derivatives.
    Author: Jiang W, Prescott M, Devenish RJ, Spiccia L, Hearn MT.
    Journal: Biotechnol Bioeng; 2009 Jul 01; 103(4):747-56. PubMed ID: 19350626.
    Abstract:
    The capabilities of a new class of immobilized (im) metal ion chelate complexes (IMCCs), derived from 1,4,7-triazacyclononane (tacn), bis(1,4,7-triazacyclononyl) ethane (dtne) and bis(1,4,7-triazacyclononyl)propane (dtnp) complexed with the borderline metal ions Cu(2+), Ni(2+), Zn(2+), Mn(2+), Co(2+), and Cr(3+), for the purification of proteins have been investigated. In particular, the binding behavior of a model protein, the C-terminal hexahistidine tagged recombinant fusion protein Schistosoma japonicum glutathione S-transferase-Saccharomyces cerevisiae mitochondrial ATP synthase delta-subunit (GST-deltaATPase-His(6)), with these new immobilized metal ion affinity chromatographic (IMAC) sorbents was compared to the properties of a conventional sorbent, derived from immobilized Ni(II)-nitrilotriacetic acid (im-Ni(2+)-NTA). Investigations using the recombinant GST-deltaATPase-His(6) and recombinant S. japonicum glutathione S-transferase (GST) lacking a hexahistidine tag have confirmed that the C-terminal tag hexahistidine residues were required for the binding process to occur with these IMAC systems. The results also confirm that recombinant fusion proteins such as GST-deltaATPase-His(6) can be isolated in high purity with these IMAC systems. Moreover, these new macrocyclic systems manifest different selectivity features as a function of pH or ionic strength when compared to the conventional, unconstrained iminodiacetic acid (IDA) or NTA chelating ligands, complexed with borderline metal ions such as Cu(2+) or Ni(2+), as IMAC systems.
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