Single-step azide introduction in proteins via an aqueous diazo transfer.
The controlled introduction of azides in proteins provides targetable handles for selective protein manipulation. We present here an efficient diazo transfer protocol that can be applied in an aqueous solution, leading to the facile introduction of azides in the side chains of lysine residues and at the N-terminus of enzymes, e.g. horseradish peroxidase (HRP) and the red fluorescent protein DsRed. The effective introduction of azides was verified by mass spectrometry, after which the azido-proteins were used in Cu(I)-catalyzed [3 + 2] cycloaddition reactions. Azido-HRP retained its catalytic activity after conjugation of a small molecule. This modified protein could also be successfully immobilized on the surface of an acetylene-covered polymersome. Azido-DsRed was coupled to an acetylene-bearing protein allowing it to act as a fluorescent label, demonstrating the wide applicability of the diazo transfer procedure.
van Dongen
Stijn F M
SF
Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525AJ Nijmegen, The Netherlands.
Teeuwen
Rosalie L M
RL
Nallani
Madhavan
M
van Berkel
Sander S
SS
Cornelissen
Jeroen J L M
JJ
Nolte
Roeland J M
RJ
van Hest
Jan C M
JC
D001386:Azides / Q000737:chemistry*; D004800:Enzymes, Immobilized / Q000737:chemistry; D006735:Horseradish Peroxidase / Q000737:chemistry; D008164:Luminescent Proteins / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D015336:Molecular Probe Techniques; D011506:Proteins / Q000737:chemistry*; D014867:Water; D000097573:Red Fluorescent Protein
D001386:Azides; D004800:Enzymes, Immobilized; D006735:Horseradish Peroxidase; D008164:Luminescent Proteins; D008239:Lysine; D011506:Proteins; D014867:Water; D000097573:Red Fluorescent Protein