Pseudo 2-fold symmetry in the copper-binding domain of arthropodan haemocyanins: Possible implications for the evolution of oxygen transport proteins

Investigation of the copper-binding centre of Panulirus interruptus haemocyanin led to the discovery of a pseudo 2-fold axis relating two helical pairs surrounding and co-ordinating the two copper ions. The pseudo 2-fold symmetry relating one helical pair, co-ordinating Cu-A, to the second helical pair co-ordinating Cu-B is quite precise with 31 equivalent Cα atoms having a root-mean-square deviation of only 1.47 Å. The 2-fold consists of a rotation of 174.6 ° and a translation parallel to the rotation axis of 0.7 Å. After superposition of the helical pairs, the two copper ions are within 1.1 Å and the three Cα atoms of the histidine ligands of Cu-A are within a root-mean-square deviation of 1.0 Å from the Cα atoms of the histidine residues co-ordinating Cu-B. Of the superimposed residues, 26% are identical in sequence. These data suggest that the current oxygen-binding centre of arthropodan haemocyanins is the result of dimerization, gene duplication and gene fusion of an ancestral mono-copper-binding helical pair. This suggestion is supported by the recent discovery that in the sequence of functional domains of molluscan haemocyanins only amino acid sequence homology with the arthropodan Cu-B helical pair has been found and no evidence for similarity with a Cu-A binding helical pair was observed. This provides strong evidence that a mono-copper-binding helical pair has been the ancestor of both the arthropodan and molluscan haemocyanins.