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Genetic Approach to Facilitate Purification of Recombinant Proteins with a Novel Metal Chelate Adsorbent

Bio/Technology volume 6, pages 1321–1325 (1988)Cite this article

Abstract

We describe a general purification method for recombinant proteins based upon the selective interaction between a poly-histidine peptide, which is fused to the protein of interest, and a novel metal chelate adsorbent. The principle of the technique is illustrated with mouse dihydrofolate reductase. DNA elements coding for adjacent histidines were fused to the mouse dihydrofolate reductase gene. Subsequent expression in E. coli resulted in the production of hybrid proteins that could be purified by immobilized metal ion affinity chromatography, followed by removal of the histidine affinity peptide with carboxypeptidase A.

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References

  1. Moks, T., Abrahmsen, L., Oesterlöf, B., Josephson, S., Oestling, M., Enfors, S.O., Persson, I., Nilsson, B. and Uhlen, M. 1987. Large-scale affinity purification of human insulin-like growth factor I from culture medium of E. coli. Bio/Technology 5:379–382.

    CAS  Google Scholar 

  2. Germio, J. and Bastia, D. 1984. Rapid purification of a cloned gene product by genetic fusion and site specific proteolysis. Proc. Nad. Acad. Sci. USA 81:4692–4696.

    Article  Google Scholar 

  3. Sassenfeld, H.M. and Brewer, S.J. 1984. A polypeptide fusion designed for the purification of recombinant proteins. Bio/Technology 2:76–81.

    CAS  Google Scholar 

  4. Hochuli, E., Döbeli, H. and Schacher, A. 1987. New metal chelate adsorbent selective for proteins and peptide containing neighbouring histidine residues. J. Chromatogr. 411:177–184.

    Article  CAS  Google Scholar 

  5. Bujard, H., Gentz, R., Lanzer, M., Stüber, D., Muller, M., Ibrahimi, I., Haeuptle, M.-T., Dobberstein, B. 1987. A T5 promoter based transcription-translation system for the analysis of proteins in vivo and in vitro. Methods in Enzymology 155:416–433.

    Article  CAS  Google Scholar 

  6. Chang, A.C.Y., Nunberg, J.H., Kaufman, R.J., Erlich, H.A., Schimke, R.T. and Cohen, S.N. 1978. Phenotypic expression in E. coli of a DNA sequence coding for mouse dihydrofolate reductase. Nature 275:617–624.

    Article  CAS  Google Scholar 

  7. Masters, J.N., Attardi, G. 1983. The nucleotide sequence of the cDNA coding for the human dihydrofolic acid reductase. Gene 21:59–63.

    Article  CAS  Google Scholar 

  8. Schwarz, E., Scherer, G., Hobom, G., Kässel, H. 1978. Nucleotide sequence of cro, cII and part of the 0 gene in phage lambda DNA. Nature 272:410–414.

    Article  CAS  Google Scholar 

  9. Marcoli, R., Iida, S., Bickle, A. 1980. The DNA sequence of an IS1-flanked transposon coding for resistance to chloramphenicol and fusidic acid. FEBS Letts 110:11–14.

    Article  CAS  Google Scholar 

  10. Brosius, J., Dull, T.J., Sleeter, D.D., Noller, H.F. 1981. Gene organization and primary structure of a ribosomal RNA operon from E. coli. J. Mol. Biol. 148:107–127.

    Article  CAS  Google Scholar 

  11. Sutcliffe, J.G. 1979. Complete nucleotide sequence of the E. coli plasmid pBR322. Cold Spring Harbor Symp. Quant. Biol. 43:77–90.

    Article  CAS  Google Scholar 

  12. Certa, U., Bannwarth, W., Stüber, D., Gentz, R., Lanzer, M., Le Grice, S., Guillot, F., Wendler, I., Hunsmann, G., Bujard, H. and Mous, J. 1986. Subregions of a conserve of part of the HIV gp 41 transmembrane protein are differentially recognized by antibodies of infected individuals. EMBO J. 5:3051–3056.

    Article  CAS  Google Scholar 

  13. Gentz, R., Certa, U., Takacs, B., Matile, H., Döbeli, H., Pink, R., Mackay, M., Bone, N. and Scaife, J.G. 1988. Major surface antigen p. 190 of Plasmodium falciparum: detection of common epitopes present in a variety of plasmodia ioslates. EMBO J. 7:225–230.

    Article  CAS  Google Scholar 

  14. Bannwarth, W., Iaiza, P. 1986. A system for the simultaneous chemical synthesis of different DNA fragments on solid support. DNA 5:413–419.

    Article  CAS  Google Scholar 

  15. Stüber, D., Bujard, H., Hochuli, E., Kocher, H.P., Kompis, I., Talmadge, K., Weibel, E.K., Winkler, F.K., Then, R.L. 1986. Enzymatic characterization of recombinant human dihydrofolate reductase produced in E. coli, p. 839–842. Chemistry and Biology of Pteridines. Cooper, B. A., Whitehead, V. M. (Eds.). Walter de Gruyter & Co. Berlin.

    Google Scholar 

  16. Laemmli, U.K. 1970. Cleavage of structural proteins during assembly of the head of bacteriophage T4 . Nature 227:680–685.

    Article  CAS  Google Scholar 

  17. Lowry, O.H. 1951. Protein measurement with folin phenol reagent. J. Biol. Chem. 193:265–275.

    CAS  Google Scholar 

  18. Baccanari, D.P., Averett, D., Briggs, C., Burchall, J. 1977. Escherichia coli dihydrofolate reductase: isolation and characterization of two isozymes. Biochemistry 16:3566–3572.

    Article  CAS  Google Scholar 

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  1. E. Hochuli: Author to whom correspondence should be sent.

Authors and Affiliations

  1. F. Hoffmann-La Roche & Co. Ltd., Central Research Units, CH-4002, Basel, Switzerland

    E. Hochuli, W. Bannwarth, H. Döbeli, R. Gentz & D. Stüber

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  1. E. Hochuli
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  2. W. Bannwarth
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  3. H. Döbeli
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  4. R. Gentz
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  5. D. Stüber
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Hochuli, E., Bannwarth, W., Döbeli, H. et al. Genetic Approach to Facilitate Purification of Recombinant Proteins with a Novel Metal Chelate Adsorbent. Nat Biotechnol 6, 1321–1325 (1988). https://doi.org/10.1038/nbt1188-1321

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