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Abstract
AbstractSynthetic metal complexes can be used as paramagnetic probes for the study of proteins and protein complexes. Herein, two transition metal NMR probes (TraNPs) are reported. TraNPs are attached through two arms to a protein to generate a pseudocontact shift (PCS) using cobalt(II), or paramagnetic relaxation enhancement (PRE) with manganese(II). The PCS analysis of TraNPs attached to three different proteins shows that the size of the anisotropic component of the magnetic susceptibility depends on the probe surroundings at the surface of the protein, contrary to what is observed for lanthanoid‐based probes. The observed PCS are relatively small, making cobalt‐based probes suitable for localized studies, such as of an active site. The obtained PREs are stronger than those obtained with nitroxide spin labels and the possibility to generate both PCS and PRE offers advantages. The properties of TraNPs in comparison with other cobalt‐based probes are discussed.
Highlights
Ever since the determination of first metalloprotein structures using paramagnetic NMR restraints,[1] it has been acknowledged that paramagnetism is a powerful tool for the study of biomolecules
Three proteins were used to characterize the paramagnetic properties of the transition metal NMR probes (TraNPs), T4 lysozyme (T4Lys) with the mutations K147C/T151C, Bacillus circulans xylanase (BCX) with mutations E78Q/T109C/T111C and ubiquitin with the mutations E24C/A28C
In T4Lys and ubiquitin, the two cysteine residues are located in an a-helix, whereas in BCX, they are in a b-strand
Summary
Can be measured with high precision. PREs only yield distance information but are strongly distance-dependent, making them exquisitely sensitive to minor states in which the nuclear-electron distance is reduced.[3]. Chemie number of cyclen derivatives have been developed for metal ions for the application in biomedicine[13] and magnetic resonance imaging (MRI),[14] for which high thermodynamic stability and kinetic inertness are required Among these reported cyclen-based complexes, there are, few successful CoII and MnII complexes, in particular for MnII, which can be oxidized by air.[15]. We report the design and synthesis of several C2symmetric cyclen derivatives, which can tightly bind the transition metal ions CoII and MnII and are stable in air and buffers These transition metal NMR probes (TraNPs) were tested using three proteins. We discuss the TraNP properties and compare them with other reported CoII tags
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