Metal-chelate support

Considerable evidence has now accumulated to show that protein are retained by a metal affinity supports according to the number of accessible histidine residues [ 140,143]. The histidine imidazole nitrogens co-ordinate metals in the unprotonated state hence in addition to the number of accessible histidines, their ionization state, that is in turn dependent upon the nature of surrounding amino acids, influence the retention of protein on metal chelate supports [143]. 

Histidine is one of the less abundant amino acids of proteins and most globular proteins contain not more that 2% histidine [146]. Considering that less than half of histidine residues of proteins are located at the surface, most proteins on the average have only about one exposed residue for every hundred amino acids. It is therefore natural that many proteins do not bind to metal chelates. Most of the work on binding and immobilization to metal chelate supports is therefore related to proteins with enriched histidines. However it has been shown that a single surface histidine may be adequate to retain a protein strongly on metal chelate supports [140]. 

Several proteins and enzymes have innate affinity in their native state for the metal chelate supports due to the presence of one or more surface histidines (Table 3). Coulet et al. [154] immobilized lactate dehydrogenase, malate dehydrogenase and alkaline phosphatase on Co2+, Zn2+, Cu2+-chelate Sepharose. All the enzymes were active in the immobilized state although the retention of the specific activity was quite low. Among the enzymes used, alkaline phosphatase retained higher activity and among the various metal chelates maximum activi-... 

Table 3. Native and affinity-tail bearing enzymes immobilized on metal chelate supports...

Anspach and Hasse [155] reported the immobilization of penicillin G amido-hydrolase from E. coli on immobilised metal chelate supports. A number of the other proteins and enzymes are also known to bind strongly to metal chelate supports including ribonuclease A and lysozyme [114]. 

On the other hand, recombinant FucA from E. coli expressed as a fusion protein with a hexa-histidine tag has been immobilized by using metal-chelate supports (Ardao et al. 2006). IMAC supports allowed FucA purification and immobilization in one step in order to obtain an immobilized FucA catalyst for aldol... 

Armisen P., Mateo C., Cortes E., Barredo J.L., Salto F., Diez B., Rodes L., Garcia J.L., Fernandez-Lafuente R. and Guisan J.M. 1999. Selective adsorption of poly-His tagged glu-taryl acylase on tailor-made metal chelate supports, J. Chromatogr. A, 848, 61. 

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... 

R3 R2 and R2 Ri gauche interactions however, for the same set of substituents, an increase in the steric requirements of either Rj or R3 will influence only one set of vicinal steric interactions (Rj R2 or R3 R2). Some support for these conclusions has been cited (eqs. [6] and [7]). These qualitative arguments may also be relevant to the observed populations of hydrogen- and nonhydrogen-bonded populations of the aldol adducts as well (see Table 1, entries K, L). Unfortunately, little detailed information exists on the solution geometries of these metal chelates. Furthermore, in many studies it is impossible to ascertain whether the aldol condensations between metal enolates and aldehydes were carried out under kinetic or thermodynamic conditions. Consequently, the importance of metal structure and enolate geometry in the definition of product stereochemistry remains ill defined. This is particularly true in the numerous studies reported on the Reformatsky reaction (20) and related variants (21). 

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]. 

Precursors of an important class of metal chelates, N,N-dialkylthiocarbamates, were used to study rearrangements in them [736], The reactions were carried out (3.282) in a microwave oven using various supports, in particular graphite, with yields 30-90%. 

In this reversed phase high performance liquid chromatographic method for neutral and cationic metal chelates with azo dyes, tetraalkylammonium salts are added to an aqueous organic mobile phase. The tetraalkylammonium in salts are dynamically coated on the reversed stationary support. As a result of the addition of tetraalkylammonium salts, the retention of the chelates is remarkably reduced. Tetrabutylammonium bromide permits rapid separation and sensitive spectrophotometric detection of the vanadium(V) chelate with 2-(8-quinolylazo)-5-(dimethylamino)-phenol, making it possible to determine trace vanadium(V). 

Eppinger, J., Nikolaides, K.R., Zhang-Presse, M. et al. (2008) Alkyl complexes of rare-earth metal centers supported by chelating l,l -diamidoferrocene ligands synthesis, structure, and application in methacrylate polymerization. Organometallics, 27, 736. 

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