Optimized His-Tag Protein Purification from Cell-Free Expression Reactions

4453MAHis-tag is the most commonly used protein fusion tag. Current available nickel purification systems are of limited use for purifying His-tagged proteins expressed in rabbit reticulocyte lysate because of the copurification of hemoglobin present in rabbit reticulocyte lysate. This often limits the downstream applications of purified proteins in fluorescence-based functional assays or protein:protein interaction studies and also decreases the amount of proteins purified.

The MagZ™ Protein Purification System, is a unique system designed for the purification of expressed His-tagged protein from rabbit reticulocyte lysate. This protein purification system has the advantage over nickel-based immobilized metal ion affinity chromatography (IMAC) because it eliminates 99.9% of hemoglobin contamination.

The three basic steps involved in His-tagged protein purification from TNT® Reticulocyte Lysate using the MagZ™ System are shown in the figure. The steps are optimized for efficient recovery of His-tagged proteins without copurification of hemoglobin. The magnetic nature of the MagZ™ Binding Particles allows purification to be performed in a single tube, simplifying and shortening the purification process to less than 30 minutes.

References citing use of the MagZ™ Protein Purification System

  1. Muik, M. et al. (2008) J. Biol. Chem. 283, 8014–22.
  2. Jurynec, M.J. et al. (2008) Proc. Natl. Acad. Sci. 105, 12485–90.
  3. Bowen, C. et.al (2007) Cancer Res. 67, 455–64.
  4. Bjorkhaug, L. et al. (2007) J. Biol. Chem. 282, 22575–64.
  5. Polesskaya, A. et al. (2007) Genes and Dev. 21, 1125–38.
  6. Godat, B. et al. (2004) Promega Notes 88, 9–12.
  7. Godat, B. et al. (2004) Promega Notes 88, 13–15.
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Gary Kobs

Strategic Marketing Manager at Promega Corporation
Gary earned his B.S. in Bacteriology, UW-Madison in 1982. From 1982–1986 he served as Research Tech at UW-Madison. From 1986 to the present Gary has been with Promega Corporation serving in many capacities including as the very first editor of Promega Notes. He was also Manager Tech Services and Training, Product Manager Restriction/Modifying Enzymes, Product Manager Protein Analysis, and is now Marketing Manager Protein Analysis.

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