Paramagnetic NMR investigations of Co(II) and Ni(II) amicyanin

The paramagnetic H-1 NMR spectra of the Co(II) and Ni(II) substituted forms of the type 1 blue copper protein (cupredoxin) amicyanin have been assigne d. This is the first such analysis of a cupredoxin, which has a distorted t etrahedral active site with the ligands provided by two histidines, a cyste ine and a methionine. The isotropic shifts of the resonances in these spect ra are compared with those of Co(II) and Ni(II) azurin. A number of interes ting similarities and differences are found. The coordination of the metal by the two equatorial histidine ligands is very similar in both proteins. T he interaction between the introduced metal and the thiolate sulfur of the equatorial cysteine ligand is enhanced in the amicyanin derivatives. Resona nces belonging to the weak axial methionine ligand exhibit much larger shif ts in the amicyanin derivatives, indicative of shorter M(II)-S(Met) distanc es. The presence of shorter axial M(II)-S(Met) and equatorial M(II)-S(Cys) distances in both Co(II) and Ni(II) amicyanin is ascribed to the absence of a second axially interacting amino acid at the active site of this cupredo xin.