A 1H NMR study of the paramagnetic active site of the CuA variant of amicyanin.
The dinuclear paramagnetic center of the CuA variant of the cupredoxin amicyanin has been investigated using 1H NMR. The hyperfine-shifted resonances have been assigned using a combination of 1D NOE difference and 2D WEFT-NOESY spectroscopy. The shifts experienced by the assigned resonances have been used to calculate hyperfine coupling constants for these protons from which the spin density distribution on the ligands at the CuA center is obtained. A comparison with published data for the paramagnetic form of wild type amicyanin highlights a number of similarities and differences between these evolutionary related sites. In both cases 50-60% of the unpaired spin density is distributed on the ligands, which in the case of the CuA center involves two cysteine and two histidine ligands. The two weak axial interactions at the CuA center carry less than 1% spin density.
Dennison
C
C
Leiden Institute of Chemistry, Leiden University, The Netherlands.
Berg
A
A
Canters
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GW
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D005075:Biological Evolution; D003300:Copper; D004926:Escherichia coli / Q000378:metabolism; D006859:Hydrogen; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008667:Metalloproteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D013855:Thiobacillus / Q000378:metabolism*
D001426:Bacterial Proteins; D008024:Ligands; D008667:Metalloproteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C017079:mauC protein, Methylobacterium extorquens; D003300:Copper; D006859:Hydrogen