The utility of pseudocontact shifts in the structure refinement of metalloproteins has been evaluated using a native, paramagnetic Cu2+ metalloprotein, plastocyanin from Anabaena variabilis (A.v.), as a model protein. First, the possibility of detecting signals of nuclei spatially close to the paramagnetic metal ion is investigated using the WEFT pulse sequence in combination with the conventional TOCSY and 1H-15N HSQC sequences. Second, the importance of the electrical charge of the metal ion for the determination of correct pseudocontact shifts from the obtained chemical shifts is evaluated. Thus, using both the Cu+ plastocyanin and Cd2+-substituted plastocyanin as the diamagnetic references, it is found that the Cd2+-substituted protein with the same electrical charge of the metal ion as the paramagnetic Cu2+ plastocyanin provides the most appropriate diamagnetic reference signals. Third, it is found that reliable pseudocontact shifts cannot be obtained from the chemical shifts of the 15N nuclei in plastocyanin, most likely because these shifts are highly dependent on even minor differences in the structure of the paramagnetic and diamagnetic proteins. Finally, the quality of the obtained 1H pseudocontact shifts, as well as the possibility of improving the accuracy of the obtained structure, is demonstrated by incorporating the shifts as restraints in a refinement of the solution structure of A.v. plastocyanin. It is found that incorporation of the pseudocontact shifts enhances the precision of the structure in regions with only few NOE restraints and improves the accuracy of the overall structure. Copyright © 2006 John Wiley & Sons, Ltd.