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Updated: Jul 18, 2026

Luminescence Resonance Energy Transfer to Study Conformational Changes in Membrane Proteins Expressed in Mammalian Cells
Published on: September 16, 2014
Long-range-distance NMR effects in a protein labeled with a lanthanide-DOTA chelate
Monica D Vlasie1, Clara Comuzzi, Adrianus M C H van den Nieuwendijk
1Leiden Institute of Chemistry, Gorlaeus Laboratories Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
Abstract:
A two-thiol reactive lanthanide-DOTA (1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid) chelate, CLaNP-3 (CLaNP=caged lanthanide NMR probe), was synthesized for the rigid attachment to cysteine groups on a protein surface, and used to obtain long-range-distance information from the {15N,1H} HSQC spectra of the protein-lanthanide complex. The DOTA ring exhibits several isomers that are in exchange; however, single resonances were observed for most amide groups in the protein, allowing determination of a single, apparent magnetic-susceptibility tensor. Pseudocontact shifts caused by Yb-containing CLaNP-3 were observed for atoms at 15-35 A from the metal. By using Gd-containing CLaNP-3, relaxation effects were observed, allowing distances up to 30 A from the paramagnetic center to be determined accurately. Similar results were obtained with a Gd-DTPA (diethylene-triaminepentaacetic acid) chelate, CLaNP-1, bound in the same bidentate manner to the protein. This study demonstrates that bidentate attachment of a paramagnetic probe enables determination of long-range distances.
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