Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Purification of recombinant proteins is often a challenging process involving several chromatographic steps that must be optimized for each target protein. Here, we developed a self-excising module allowing single-step affinity chromatography purification of untagged recombinant proteins. It consists of a 250-residue-long self-processing module of the Neisseria meningitidis FrpC protein with a C-terminal affinity tag. The N terminus of the module is fused to the C terminus of a target protein of interest. Upon binding of the fusion protein to an affinity matrix from cell lysate and washing out contaminating proteins, site-specific cleavage of the Asp-Pro bond linking the target protein to the self-excising module is induced by calcium ions. This results in the release of the target protein with only a single aspartic acid residue added at the C terminus, while the self-excising affinity module remains trapped on the affinity matrix. The system was successfully tested with several target proteins, including glutathione-S-transferase, maltose-binding protein, beta-galactosidase, chloramphenicol acetyltransferase, and adenylate cyclase, and two different affinity tags, chitin-binding domain or poly-His. Moreover, it was demonstrated that it can be applied as an alternative to two currently existing systems, based on the self-splicing intein of Saccharomyces cerevisiae and sortase A of Staphylococcus aureus.
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PMID:Single-step affinity purification of recombinant proteins using a self-excising module from Neisseria meningitidis FrpC. 1866 6

A new method is described for facile synthesis of metal-chelating magnetic nanoparticles by simply mixing iron oxide nanoparticles with a bifunctional organophosphorus compound, N-(phosphonomethyl)iminodiacetic acid (PM-IDA), in aqueous solution. On charging with nickel ions, the PM-IDA functionalized iron oxide nanoparticles exhibited high His-tag protein binding capacity (0.21 and 0.58 mg/mg for His-tagged green fluorescent protein and chloramphenicol acetyltransferase, respectively) and were successfully used to purify these proteins from bacterial cell extracts to high purity in a single step. Although other synthetic schemes for metal-chelating magnetic nanoparticles have been reported, the method described here is markedly simpler and involves only low-cost reagents.
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PMID:Facile synthesis of metal-chelating magnetic nanoparticles by exploiting organophosphorus coupling. 2085 Apr 8

Specific tags with defined amino acid residues are widely used to purify or probe target proteins. Interestingly, the tagging system occasionally results in an increase of the recombinant protein expression in vivo. Here, we systematically examined this phenomenon using a poly-histidine (His)-tag fused to N- or C-terminal region of green, red, and blue fluorescent proteins by quantification and uneven distribution in cytoplasm of Escherichia coli. This effect was further supported by the distinct over-expression of several unrelated proteins, such as esterase, neopullulanase, and chloramphenicol acetyltransferase, tagging with the same tag. These results suggest that a poly-His-tag placed at N-terminal region can induce over-expression of recombinant protein via subcellular uneven distribution in vivo.
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PMID:Over-expression of recombinant proteins with N-terminal His-tag via subcellular uneven distribution in Escherichia coli. 2622 Sep 49


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