Metal-interaction affinity chromatography

Metal-chelate affinity chromatography is a powerful purification technique whereby proteins or other molecules can be separated based upon their ability to form coordination complexes with immobilized metal ions (Porath et al., 1975 Lonnerdal and Keen, 1982 Porath and Belew, 1983 Porath and Olin, 1983 Sulkowski, 1985 Kagedal, 1989). The metal ions are stabilized on a matrix through the use of chelating compounds which usually have multivalent points of interaction with the metal atoms. To form useful affinity supports, these metal ion complexes must have some free or weakly associated and exchangeable coordination sites. These exchangeable sites then can form complexes with coordination sites on proteins or other molecules. Substances that are able to interact with the immobilized metals will bind and be retained on... 

In 1970s, first application of metal-chelate affinity chromatography which is later named as "immobilized-metal (ion) affinity chromatography (IMAC) was perfomed. Metal-chelate chromatography technique exploits selective interactions and affinity between transition metal immobilized on a solid support (resin) via a metal chelator and amino acid residues which act as electron donors in the protein of interest [25-26]. As well as aromatic and heterocyclic compounds, proteins such as histidine, tyrosine, tyriptophane and phenylalanine posses affinity to transition metals which form complexes with compounds rich in electrons [25,27]. 

Immobilized metal-ion affinity chromatography (IMAC) is a collective term that includes all kinds of affinity chromatography, where metal atoms or ions immobilized on polymer cause or dominate the interaction at the sorption site. 

A variety of micropellicular packing materials has been developed for the analysis of both small and large molecules by various HPLC modes, including ion exchange (lEC), metal interaction (MIC), reversed phase (RPC) [4], and affinity chromatography (AC) [5]. Besides analytical applications, other possible utilization of micropellicular stationary phases includes fundamental kinetic and thermodynamic studies of the retention mechanisms on a well-defined surface. Nevertheless, a relatively limited variety of micropellicular... 

Affinity separation of proteins in immobilized metal ion affinity chromatography (IMAC). The method is based on the fact that some protein molecules, having specific residues on the surface, can form complexes with metal ions immobilized on the sorbent matrix. Retention is determined by the coordination interactions forming the complexes. Immobilized metal ion affinity chromatography can be performed under very mild, nondenaturing conditions with extremely high selectivity it is particularly suitable for preparative group fractionation of complex extracts and bio-fluids. 

Immobilized metal ion affinity chromatography (IMAC) rehes on the ability of certain amino acid side chains to form coordinate bonds with immobilized metal ion complexes [6,181-186]. The adsorption of proteins mainly takes place through interactions with the imidazole ring of histidines. For other amino acids with electron donor atoms in their side chains, binding tends to be weak. Cysteines in natural proteins, are rarely available in an appropriate reduced form for binding to chelated metal ions. 

Metal-chelate affinity chromatography was introduced as a specific method for fractionation of proteins by Porath et al., in 1975. The principle of this type of chromatographic method is that certain amino acid residues, such as histidine, cysteine, lysine, tryptophan, aspartic acid, glutamic acid, or phosphorylated amino acids, which are accessible on the protein surface, can interact through non-bonding lone-pair electron coordination with some metal ions. Metal cations Cu, Ni, ... 

Metal interaction chromatography is an HPLC technique that can separate many biopolymers because of their differential ability to form complexes with metal ions [1]. It employs a stationary phase with an appropriate metal immobilized via chelating functions bound to the surface. Retention and separation of the sample components occur largely by their interaction with the chelated metal. Although immobilized metal affinity chromatography (IMAC) is a common name for the technique, it is called metal interaction chromatography (MIC) in this book to conform to the nomenclature used for the other interactive chromatographic methods for biopolymer separation by HPLC. 

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