Details

Autor(en) / Beteiligte

• Kodal, Anne Louise B.
• Rosen, Christian B.
• Mortensen, Michael R.
• Tørring, Thomas
• Gothelf, Kurt V.

Titel

DNA-Templated Introduction of an Aldehyde Handle in Proteins

Ist Teil von

• Chembiochem : a European journal of chemical biology, 2016-07, Vol.17 (14), p.1338-1342

Ort / Verlag

Germany: Blackwell Publishing Ltd

Erscheinungsjahr

2016

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Many medical and biotechnological applications rely on protein labeling, but a key challenge is the production of homogeneous and site‐specific conjugates. This can rarely be achieved by simple residue‐specific random labeling, but generally requires genetic engineering. Using site‐selective DNA‐templated reductive amination, we created DNA–protein conjugates with control over labeling stoichiometry and without genetic engineering. A guiding DNA strand with a metal‐binding functionality facilitates site‐selectivity by directing the coupling of a second reactive DNA strand in the vicinity of a protein metal‐binding site. We demonstrate DNA‐templated reductive amination for His6‐tagged proteins and metal‐binding proteins, including IgG1 antibodies. We also used a cleavable linker between the DNA and the protein to remove the DNA and introduce a single aldehyde on the protein. This functions as a handle for further modifications with desired labels. In addition to directing the aldehyde positioning, the DNA provides a straightforward route for purification between reaction steps. Site‐selective labeling of metal‐binding proteins is achieved by metal‐coordination and DNA‐templated reductive amination of a DNA‐tethered aldehyde with lysine at the surface of the protein, followed by oxidative cleavage of a diol spacer in order to remove the DNA and form the aldehyde handle at the protein.