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

Bio/Technologyvolume 13pages504506 (1995) | Download Citation

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Abstract

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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Affiliations

  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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https://doi.org/10.1038/nbt0595-504

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