title: A caged lanthanide complex as a paramagnetic shift agent for protein NMR.
abstract: A lanthanide complex, named CLaNP (caged lanthanide NMR probe) has been developed for the characterisation of proteins by paramagnetic NMR spectroscopy. The probe consists of a lanthanide chelated by a derivative of DTPA (diethylenetriaminepentaacetic acid) with two thiol reactive functional groups. The CLaNP molecule is attached to a protein by two engineered, surface-exposed, Cys residues in a bidentate manner. This drastically limits the dynamics of the metal relative to the protein and enables measurements of pseudocontact shifts. NMR spectroscopy experiments on a diamagnetic control and the crystal structure of the probe-protein complex demonstrate that the protein structure is not affected by probe attachment. The probe is able to induce pseudocontact shifts to at least 40 A from the metal and causes residual dipolar couplings due to alignment at a high magnetic field. The molecule exists in several isomeric forms with different paramagnetic tensors; this provides a fast way to obtain long-range distance restraints.
authors.affiliation: Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
authors.forename: Miguel
authors.initials: M
authors.lastname: Prudêncio
authors.forename: Jan
authors.initials: J
authors.lastname: Rohovec
authors.forename: Joop A
authors.initials: JA
authors.lastname: Peters
authors.forename: Elitza
authors.initials: E
authors.lastname: Tocheva
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Boulanger
authors.forename: Michael E P
authors.initials: ME
authors.lastname: Murphy
authors.forename: Hermen-Jan
authors.initials: HJ
authors.lastname: Hupkes
authors.forename: Walter
authors.initials: W
authors.lastname: Kosters
authors.forename: Antonietta
authors.initials: A
authors.lastname: Impagliazzo
authors.forename: Marcellus
authors.initials: M
authors.lastname: Ubbink
mesh_terms: D001400:Azurin / Q000031:analogs & derivatives* / Q000737:chemistry; D018360:Crystallography, X-Ray; D028581:Lanthanoid Series Elements / Q000138:chemical synthesis / Q000737:chemistry*; D013058:Mass Spectrometry; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D004369:Pentetic Acid / Q000737:chemistry*
chemical_list: D028581:Lanthanoid Series Elements; D001400:Azurin; D004369:Pentetic Acid