The 800-MHz 1H NMR spectra of oxidized plastocyanin from spinach are here reported. All hyperfine-shifted signals have been assigned through saturation transfer with the reduced diamagnetic species. To detect the copper(II)-bound cysteine β-CH2 signals, a technique has been applied which is based on irradiation of regions where such signals are expected but not detected, and the corresponding saturation transfer on the reduced species is observed. At the end, a full spectrum is reconstructed which permits, for the first time, the complete 1H NMR signal assignment of an oxidized blue copper protein. These data are discussed in terms of the factors affecting the line width as related to the electronic and geometric structure of the metal center. A Karplus-type relationship is proposed between the contact shift of the Cys-84 β-CH2 protons and the Cu−S−C−Hβ dihedral angle.

Bacillus pasteurii UreE (BpUreE) is a putative chaperone assisting the insertion of Ni 2؉ ions in the active site of urease. The x-ray structure of the protein has been determined for two crystal forms, at 1.7 and 1.85 Å resolution, using SIRAS phases derived from a Hg 2؉ -derivative. BpUreE is composed of distinct N-and Cterminal domains, connected by a short flexible linker. The structure reveals the topology of an elongated homodimer, formed by interaction of the two C-terminal domains through hydrophobic interactions. A single Zn 2؉ ion bound to four conserved His-100 residues, one from each monomer, connects two dimers resulting in a tetrameric BpUreE known to be formed in concentrated solutions. The Zn 2؉ ion can be replaced by Ni 2؉ as shown by anomalous difference maps obtained on a crystal of BpUreE soaked in a solution containing NiCl 2 . A large hydrophobic patch surrounding the metal ion site is surface-exposed in the biologically relevant dimer. The BpUreE structure represents the first for this class of proteins and suggests a possible role for UreE in the urease nickel-center assembly.The molecular details of the mechanism by which specific metal co-factors are transported in the cell and sorted into the correct metalloenzyme, among the many biological systems that employ transition metal ions for their function without being involved in alternative adventitious and possibly poisonous binding, are still rather unclear. The role of a new class of soluble metal-binding proteins, known as "metallochaperones

The NMR solution structure of oxidized plastocyanin from the cyanobacterium Synechocystis PCC6803 is here reported. The protein contains paramagnetic copper(II), whose electronic relaxation times are quite unfavorable for NMR solution studies. The structure has been solved on the basis of 1041 meaningful NOESY cross-peaks, 18 1D NOEs, 26 T(1) values, 96 dihedral angle constraints, and 18 H-bonds. The detection of broad hyperfine-shifted signals and their full assignment allowed the identification of the copper(II) ligands and the determination of the Cu-S-C-H dihedral angle for the coordinated cysteine. The global root-mean-square deviation from the mean structure for the solution structure family is 0.72 +/- 0.14 and 1.16 +/- 0.17 A for backbone and heavy atoms, respectively. The structure is overall quite satisfactory and represents a breakthrough, in that it includes paramagnetic copper proteins among the metalloproteins for which solution structures can be afforded. The comparison with the available X-ray structure of a triple mutant is also performed.