Immobilized metal-ion affinity chromatography: Imidazole proton pump and chromatographic sequelae. II. Chromatographic sequelae

Abstract

Isocratic elution with imidazole of some protein models from a chelating gel, CSFF-IDA-M(II), resulted in their desorption owing to the low pH(6-->4) of the mobile, phase rather than to the imidazole itself (imidazole-generated fall in pH; proton pump). Gradient elution with imidazole was best accomplished when the chelating gel was initially converted into its imidazole complex, CSFF-IDA-M(II)Im. The exploitation of the imidazole-generated proton pump in the IMAC of proteins may enhance the versatility of this type of chromatography.

Production, purification and diagnostic application of filarial recombinant protein WbSXP-1 expressed in salt inducible Escherichia coli

“…The puriWcation of any protein by chromatography and number of steps required varies and depends on the nature of protein itself [18,19]. Multi-step puriWcation strategies reduce the recovery of the target protein though the purity may improve tremendously [28]. The purity and quality of rWbSXP-1 was observed to be very critical in performance of the diagnostic kit.…”

Immobilized metal-ion affinity systems for recovery and structure–function studies of proteins at molecular, supramolecular, and cellular levels

“…In this way, IMAC can serve as a sensitive tool for revealing histidine topography on the surface of protein. Some interesting examples of using IMAC for purification of proteins with exposed, native surface histidine residues include human serum proteins [1,28], interferon [7], lactoferrin and myoglobin [29], tissue plasminogen activator [30], antibodies [31,32], and yeast alcohol dehydrogenase [33]. Purification of IgG from human serum has been carried out using metal chelate methacrylate monolith (CIM disk, unpublished) (Fig.…”

Molecular Dynamics Simulations of Metal Ion Binding to the His-tag Motif

“…Imidazole molecules compete with protein molecules for metal ion binding and thus displace bound proteins. 17 In general, imidazole is applied in concentrations higher than 0.1 M. [18][19] Accordingly, IMAC technology appears as one of the major preparative methodologies used for recombinant protein purification.…”