Whilst, the UV-vis spectroscopy and ICP-MS studies showed copper binding to the ArMs (see ESIa), they give no information about the precise environment of the copper. We therefore looked to X-ray crystallography to obtain structural understanding of the newly created active sites within the metalloproteins. Crystals of SCP_Q111CBpy and SCP_Q111BpyAla were soaked in solutions of Cu(II) ions and their structures determined by X-ray crystallography (Figure and). Triton-X-100 was needed for crystallisation, similar to wt SCP-2L (PDB ID: 1IKT) ,the apolar tunnel is occupied by Triton X-100 in the structures of Cu(II)-bound SCP_Q111CBpy and SCP_Q111BpyAla (Figure ). The structure of Cu(II)-bound SCP_Q111CBpy was determined to 1.52 Å resolution, and clearly confirmed the incorporation of 2,2'-bipyridine at C111, which is positioned at the centre of the C-terminal helix,5. The bipyridine rings are sandwiched in a parallel fashion between the amide side-chain atoms of Q108 (one turn up on -helix 5) and Q90 on -helix 4 (Figure ). The outer and inner pyridine rings are 3.4 Å from Q108 and Q90 respectively, a distance that favours formation of stabilising aromatic -amino electrostatic interactions. Helix 5 is slightly shifted relative to its position in the wt SCP_2L structure (PDB ID 1IKT) (Figure in ESIa) and is more flexible than the remainder of the structure, with B-factors ranging from 52-96 Å 2 from the N-to the C-terminus of the helix (N103-L120), compared to 75 Å 2 for the protein overall. The B-factor of the Bpy moiety is 92 Å 2 , suggesting flexibility in its position. Additional electron density was observed in the 2Fo-Fc map around the solvent-exposed face of the Bpy moiety, and a single peak in an anomalous difference map confirmed this is a bound Cu(II) ion (Figure ). The Cu(II) atom lies in the same plane as the two pyridine rings and is 2 Å from the two pyridine ring nitrogens, and thus we assume that the Cu(II) ion adopts an octahedral coordination geometry. The high B-factor (108 Å) of the Cu(II) is consistent with its position in the flexible -helix 5. Only one water molecule could be modelled around the Cu(II) ion, but the positions of Q108 and Q90 suggest that their amide side-chain atoms may play a role in coordinating the other (unmodelled) waters in the coordination sphere.