Cleavage of recombinant proteins at poly-His sequences by Co(II) and Cu(II)

Research output: Contribution to journalArticleScientificpeer-review

Researchers

Research units

  • VTT Technical Research Centre of Finland

Abstract

Improved ways to cleave peptide chains at engineered sites easily and specifically would form useful tools for biochemical research. Uses of such methods include the activation or inactivation of enzymes or the removal of tags for enhancement of recombinant protein expression or tags used for purification
of recombinant proteins. In this work we show by gel electrophoresis and mass spectroscopy that salts of Co(II) and Cu(II) can be used to cleave fusion proteins specifically at sites where sequences of His residues have been introduced by protein engineering. The His residues could be either consecutive or spaced with other amino acids in between. The cleavage reaction required the presence of low concentrations of ascorbate and in the case of Cu(II) also hydrogen peroxide. The amount of metal ions required for cleavage was very low; in the case of Cu(II) only one to two molar equivalents of Cu(II) to protein was required. In the case of Co(II), 10 molar equivalents gave optimal cleavage. The reaction occurred within minutes, at a wide pH range, and efficiently at temperatures ranging from 0°C to 70°C. The work described here can also have implications for understanding protein stability in vitro and in vivo.

Details

Original languageEnglish
Pages (from-to)1751-1761
Number of pages11
JournalProtein Science
Volume16
Issue number8
Publication statusPublished - Aug 2007
MoE publication typeA1 Journal article-refereed

    Research areas

  • artificial protease, Co(II), Cu(II), His tag, protein cleavage, protein stability

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