THE ENVIRONMENT OF MULTINUCLEAR COPPER METAL LINKAGES IN PROTEIN STRUCTURES

The extended environment, particularly of the second shell about multi nuclear copper centers in protein tertiary structures, can be describe d in terms of polarity, hydrophobicity, secondary structures, solvent accessibility, H-bonding networks, and the presence of statistically s ignificant residue clusters (e.g., histidine-acidic, histidine-cystein e-methionine). Six protein structures with a multinuclear copper core are examined: ascorbate oxidase [oxidized (1aoz - Protein Data Bank (P DB) code) and reduced form (1aso)], ceruloplasmin (hCP - PDB number un assigned), hemocyanin (oxidized 1oxy and deoxygenated forms 1lla), and nitrite reductase (2afn). Analysis of these descriptors for these cop per-containing proteins reveals both known and also previously undescr ibed and possible functionally important features. The novel features are firstly that the trinuclear copper complex in ascorbate oxidase an d in ceruloplasmin are similar in that each structure is coordinated i n the same 3D orientation by four histidine pairs in the same primary sequence arrangement. By contrast, the second shell of (1aoz) is predo minantly polar augmented with three water molecules, whereas the secon d shell of (hCP) is predominantly hydrophobic. Both structures possess a histidine-acidic cluster overlapping the trinuclear copper environm ent and extending to a nearby (about 13 Angstrom away) blue copper (ty pe I) center. While the acidic residues in this cluster are not copper ligands, in (hCP) they putatively bind iron ions as part of the enzym e ferroxidase activity. Ceruloplasmin also contains two other separate d type I copper ions, and there is a distinctive cysteine-histidine-me thionine residue cluster connecting these two separated copper environ ments. We also identified in ceruloplasmin a significant mixed-charge cluster on the protein surface distant from the several copper ions wh ich may play a role in protein-protein interaction. The second shell o f the oxidized dicopper hemocyanin is more hydrophilic than that of th e reduced form, the latter being totally buried. The exclusive histidi ne-acidic and the methionine-cysteine-histidine clusters in nitrite re ductase traverse the environment of the Cu (type I) and the nearby (12 .7 Angstrom) functionally linked Cu (type II). Analysis of repeats in ceruloplasmin reveals a triple repeat consonant with the three copper type I occurrences but not consistent with the ligation patterns of th e trinuclear copper complex, suggesting the possibility of alternative coordination ligand choices and/or alternative structural conformatio nal outcomes.