Solution Structure of the Type 1 Blue Copper Protein Amicyanin from Thiobacillus versutus

Abstract

A three-dimensional solution structure of amicyanin from Thiobacillus versutus has been determined by distance geometry and restrained molecular dynamics. A total of 984 experimentally derived constraints were used for the final refinement (881 distance constraints and 103 dihedral angle constraints). Stereospecific assignments were made for 17 prochiral β-methylene protons (33%) and the methyl groups of eight valine residues. Fourteen structures were selected to represent the solution structure. They show an average pairwise backbone root-mean-square deviation of 1·19 Å. The overall structure can be described as a β-sandwich, built up of nine β-strands. The copper atom is located between three loops on one end of the molecule. Two of these loops contribute the copper ligands. His54 is on the loop between β-strands 4 and 5. The other three ligands, Cys93, His96 and Met99, are located evenly spaced on the loop between β-strands 8 and 9. This loop is folded in two consecutive type 1 turns with His96 as the donor and acceptor of the NH i-CO i- 3 hydrogen bonds. The folding is reminiscent of the general cupredoxin fold. Considerably different are the large 21 residue N-terminal extension, that is unique to amicyanin and forms an extra β-strand (strand 1), and the region between β-strands 5 and 7. The partly surface-exposed copper ligand His96 is surrounded by a hydrophobic patch consisting of seven residues.