Application of immobilized metal ion chelate complexes as pseudocation exchange adsorbents for protein separation.
The interactions of horse muscle myoglobin (MYO), tuna heart cytochrome c (CYT), and hen egg white lysozyme (LYS) with three different immobilized metal ion affinity (IMAC) adsorbents involving the chelated complexes of the hard Lewis metal ions Al3+, Ca2+, Fe3+, and Yb3+ and the borderline Lewis metal ion Cu2+ have been investigated in the presence of low- and high-ionic strength buffers and at two different pH values. In contrast to the selectivity behavior noted with buffers of high ionic strength, with low-ionic strength buffers, these three proteins interact with the hard metal ion IMAC adsorbents in a manner more characteristic of cation exchange behavior, although in contrast to the cation exchange chromatography of these proteins, as the pH value of the elution buffer was increased, the retention also increased. The selectivity differences observed under these conditions appear to be due to the formation of hydrolytic complexes of these immobilized metal ion chelate systems involving a change in the coordination geometry of the im-M(n+)-chelate at higher pH values. The experimental observations have been evaluated in terms of the effective charge on the immobilized metal ion chelate complex and the charge characteristics of the specific proteins.
Zachariou
M
M
Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Hearn
M T
MT
D000327:Adsorption; D000818:Animals; D002412:Cations; D002614:Chelating Agents; D002645:Chickens; D002846:Chromatography, Affinity / Q000379:methods*; D003574:Cytochrome c Group / Q000302:isolation & purification* / Q000378:metabolism; D005260:Female; D006736:Horses; D008670:Metals; D009113:Muramidase / Q000302:isolation & purification* / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D009211:Myoglobin / Q000302:isolation & purification* / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification*; D014413:Tuna
D002412:Cations; D002614:Chelating Agents; D003574:Cytochrome c Group; D008670:Metals; D009211:Myoglobin; D011506:Proteins; D009113:Muramidase