Structural analysis and classification of native proteins from E. coli commonly co-purified by immobilised metal affinity chromatography.
Immobilised metal affinity chromatography (IMAC) is the most widely used technique for single-step purification of recombinant proteins. However, despite its use in the purification of heterologue proteins in the eubacteria Escherichia coli for decades, the presence of native E. coli proteins that exhibit a high affinity for divalent cations such as nickel, cobalt or copper has remained problematic. This is of particular relevance when recombinant molecules are not expressed at high levels or when their overexpression induces that of native bacterial proteins due to pleiotropism and/or in response to stress conditions. Identification of such contaminating proteins is clearly relevant to those involved in the purification of histidine-tagged proteins either at small/medium scale or in high-throughput processes. The work presented here reviews the native proteins from E. coli most commonly co-purified by IMAC, including Fur, Crp, ArgE, SlyD, GlmS, GlgA, ODO1, ODO2, YadF and YfbG. The binding of these proteins to metal-chelating resins can mostly be explained by their native metal-binding functions or their possession of surface clusters of histidine residues. However, some proteins fall outside these categories, implying that a further class of interactions may account for their ability to co-purify with histidine-tagged proteins. We propose a classification of these E. coli native proteins based on their physicochemical, structural and functional properties.
Bolanos-Garcia
Victor Martin
VM
Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, England. victor@cryst.bioc.cam.ac.uk
Davies
Owen Richard
OR
D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D008670:Metals / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding
D029968:Escherichia coli Proteins; D008670:Metals