Journal
CHEMISTRY-A EUROPEAN JOURNAL
Volume 21, Issue 13, Pages 5084-5092Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/chem.201406274
Keywords
click chemistry; lanthanide-binding tags; NMR spectroscopy; p-azido-L-phenylalanine; pseudocontact shifts
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Funding
- Australian Research Council
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Pseudocontact shifts (PCS) induced by paramagnetic lanthanide ions provide unique long-range structural information in nuclear magnetic resonance (NMR) spectra, but the site-specific attachment of lanthanide tags to proteins remains a challenge. Here we incorporated p-azidophenylalanine (AzF) site-specifically into the proteins ubiquitin and GB1, and ligated the AzF residue with alkyne derivatives of small nitrilotriacetic acid and iminodiacetic acid tags using the Cu-I-catalysed click reaction. These tags form lanthanide complexes with no or only a small net charge and produced sizeable PCSs with paramagnetic lanthanide ions in all mutants tested. The PCSs were readily fitted by single magnetic susceptibility anisotropy tensors. Protein precipitation during the click reaction was greatly alleviated by the presence of 150 mm NaCl.
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