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Improved N-terminal Processing of Recombinant Proteins Synthesized in Escherichia coli

Bio/Technologyvolume 13pages504506 (1995) | Download Citation



Preparations of rHMfA (recombinant histone A from M ethanothermus f ervidus) synthesized in E. coli by the heterologous expression of the hmfA gene were found to contain a mixture of rHMf A molecules, 40% that retained the N-terminal formyl-methionyl residue (f-met-rHMfA), 50% that lacked the formyl moiety but retained the methionyl residue (met-rHMfA), and only 10% that had lost both components of the protein synthesis initiating amino acid residue and therefore had the same N-terminal sequence as native HMf A molecules synthesized in Mt. fervidus. Expression of the hmfA gene in E. coli cells grown in the presence of trimethoprim and thymidine, coupled with the concurrent over-expression of a methionine aminopeptidase-encoding map gene, has been shown to overcome this N-terminal heterogeneity problem and to result in rHMf A preparations in which >85% of the molecules have the fully processed, native N-terminal sequence. This procedure should be generally useful for ensuring N-terminal processing of recombinant proteins synthesized in E. coli.

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  1. 1

    Sherman, F., Stewart, J.W. and Tsunasawa, S. 1985. Methionine or not methionine at the beginning of a protein. BioEssays 3: 27–31.

  2. 2

    Adams, J.M. 1968. On the release of the formyl group from nascent proteins. J. Mol. Biol. 33: 571–589.

  3. 3

    Mazel, D., Pochet, S. and Marlière, P. 1994. Genetic characterization of polypeptide deformylase, a distinctive enzyme of eubacterial translation. EMBO J. 13: 914–923.

  4. 4

    Sandman, K., Grayling, R.A., Dobrinski, B., Lurz, R. and Reeve, J.N. 1994. Growth phase dependent synthesis of histones in the archaeon Methanothermus fervidus. Proc. Natl. Acad. Sci. USA 91: 12624–12628.

  5. 5

    Zillig, W., Palm, P., Reiter, W-D., Gropp, F., Püller, G. and Klenk, H-P. 1988. Comparative evaluation of gene expression in archaebacteria. Eur. J. Biochem. 173: 473–482.

  6. 6

    Sandman, K., Krzycki, J.A., Dobrinski, B., Lurz, R. and Reeve, J.N. 1990. HMf, a DNA-binding protein isolated from the hyperthermophilic archaeon Methanothermus fervidus, is most closely related to histones. Proc. Natl. Acad. Sci. USA 87: 5780–5791.

  7. 7

    LeGendre, N., Mansfield, M., Weiss, A. and Matsudaira, P. 1993. Purification of proteins and peptides by SDS-PAGE, p. 71–101. In: Practical Guide to Protein and Peptide Purification for Microsequencing. Matsudaira, P. (Ed.). Academic Press, San Diego, CA.

  8. 8

    Ben-Bassat, A., Bauer, K., Chang, S.-Y., Myambo, K., Boosman, A. and Chang, S. 1987. Processing of the initiation methionine from proteins: properties of the Escherichia coli methionine aminopeptidase and its gene structure. J. Bacteriol. 169: 751–757.

  9. 9

    Tabassum, R., Sandman, K.M. and Reeve, J.N. 1992. HMt, a histonerelated protein from Methanobacterium thermoautotrophicum ΔH. J. Bacteriol. 174: 7890–7895.

  10. 10

    Smith, B.J. 1984. Acetic acid-urea polyacrylamide gel electrophoresis of proteins, p. 63–73. In: Methods of Molecular Biology, Volume 1, Proteins. Walker, J. M. (Ed.). Humana Press, Clifton, NJ.

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  1. Department of Microbiology, The Ohio State University, 484 W. 12th Avenue, Columbus, OH, 43210

    • Kathleen Sandman
    • , Rowan A. Grayling
    •  & John N. Reeve


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Correspondence to John N. Reeve.

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