01-Jul-2008 - Royal Society of Chemistry (RSC)

Chemical tag to track proteins in cells

Native Chemical Ligation (NCL) is a reaction that takes place between a molecule containing a thioester group and a polypeptide that has a cysteine amino acid at its N-terminal. The reaction takes place in aqueous solution and mild conditions and is completely specific which ensures that a chemical tag is introduced at a single site.

NCL has inspired Robin Leatherbarrow and colleagues at Imperial College London, UK, to chemically tag a protein molecule using the enzyme protease 3Cpro from the Foot-and-Mouth Disease Virus. 3Cpro hydrolyses proteins at a specific recognition sequence. By introducing this recognition sequence followed by a cysteine residue, and then the remainder of the protein, Leatherbarrows' team has found that the 3Cpro efficiently cuts the material to leave an N-terminal cysteine at the end of the desired protein. The target protein can then be tracked in cells by using a fluorescent label.

The work was carried out as part of the EPSRC-funded Doctoral Training Centre programme of the Chemical Biology centre at Imperial College London which seeks answers to biological questions using physical sciences tools. 'One challenge of studying complex protein mixtures is to label proteins in such a way that the biological function is not compromised' says Leatherbarrow.

'Our NCL/3Cpro strategy is a useful addition to the Chemical Biologist's toolbox, providing facile, selective and generic labelling sytems', says Leatherbarrow. The generic nature of the technique suggests that it can be applied to a range of labelling problems and could lead to semi-synthetic proteins being developed.

Original publication. Robin J. Leatherbarrow et al., Chem. Commun., 2008

Facts, background information, dossiers
  • proteins
  • solution
  • Enzyme
  • chemical biology
  • ?-amino acids
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