Electron Tunneling in Single Crystals of Pseudomonas aeruginosa Azurins

Abstract

Rates of reduction of Os(III), Ru(III), and Re(I)^* by Cu(I) in His83-modified Pseudomonas aeruginosa azurins (M−Cu distance ∼17 Å) have been measured in single crystals, where protein conformation and surface solvation are precisely defined by high-resolution X-ray structure determinations:  1.7(8) × 10^6 s^(-1) (298 K), 1.8(8) × 10^6 s^(-1) (140 K), [Ru(bpy)_2(im)^(3+)-]; 3.0(15) × 10^6 s^(-1) (298 K), [Ru(tpy)(bpy)^(3+)-]; 3.0(15) × 10^6 s^(-1) (298 K), [Ru(tpy)(phen)^(3+)-]; 9.0(50) × 10^2 s^(-1) (298 K), [Os(bpy)2(im)^(3+)-]; 4.4(20) × 10^6 s^(-1) (298 K), [Re(CO)_3(phen)^(+*)] (bpy = 2,2'-bipyridine; im = imidazole; tpy = 2,2':6',2' '-terpyridine; phen = 1,10-phenanthroline). The time constants for electron tunneling in crystals are roughly the same as those measured in solution, indicating very similar protein structures in the two states. High-resolution structures of the oxidized (1.5 Å) and reduced (1.4 Å) states of Ru(II)(tpy)(phen)(His83)Az establish that very small changes in copper coordination accompany reduction but reveal a shorter axial interaction between copper and the Gly45 peptide carbonyl oxygen [2.6 Å for Cu(II)] than had been recognized previously. Although Ru(bpy)_2(im)(His83)Az is less solvated in the crystal, the reorganization energy for Cu(I) → Ru(III) electron transfer falls in the range (0.6−0.8 eV) determined experimentally for the reaction in solution. Our work suggests that outer-sphere protein reorganization is the dominant activation component required for electron tunneling.

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