Part:BBa_K5357002
6 x His Tag
Usage and Biology
A 6 x Histidine tag is can be fused toa protein, via the N- or C terminal, in order to purify it. It is comprised of 6 consecutive Histidine amino acids. The method used to purify the His tag-containing protein is called IMAC, or immobilised metal affinity chromatography. This is where divalent metal ions such as Nickel or Cobalt are chelated to a resin in a column. The lysate containing the protein with the His tag is flowed through the column. The metal ions are used to bind and immobilise the protein via its His tag, exploiting the high affinity the His tag has to these metal ions. The column is then washed with a buffer to remove unbound molecules, leaving the bound protein containing the His tag. This protein can then be eluted (and thus purified) by adding imidazole, which competes with the Histidine tag for the metal ions, thus causing the His-tagged protein to fall out of the column.
Design Notes
codon optimised for E. coli. We ensured that the sequence did not contain an excessive number of consecutive identical codons to avoid ribosomal stalling caused by a shortage of the corresponding cognate aminoacyl-tRNA. We therefore made sure that multiple synonymous codons were used for these repetitive sequences.
References
Schafer F, Romer U, Emmerlich M, Blumer J, Lubenow H, Steinert K. Automated high-throughput purification of 6xHis-tagged proteins. J Biomol Tech. 2002;13(3):131-42.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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