Search for: All records

Abstract The key to type 1 copper (T1Cu) function lies in the fine tuning of the Cu^II/Ireduction potential (E°′_T1Cu) to match those of its redox partners, enabling efficient electron transfer in a wide range of biological systems. While the secondary coordination sphere (SCS) effects have been used to tuneE°′_T1Cuin azurin over a wide range, these principles are yet to be generalized to other T1Cu‐containing proteins to tune catalytic properties. To this end, we have examined the effects of Y229F, V290N and S292F mutations around the T1Cu of small laccase (SLAC) fromStreptomyces coelicolorto match the highE°′_T1Cuof fungal laccases. Using ultraviolet‐visible absorption and electron paramagnetic resonance spectroscopies, together with X‐ray crystallography and redox titrations, we have probed the influence of SCS mutations on the T1Cu and correspondingE°′_T1Cu. While minimal and smallE°′_T1Cuincreases are observed in Y229F‐ and S292F‐SLAC, the V290N mutant exhibits a majorE°′_T1Cuincrease. Moreover, the influence of these mutations onE°′_T1Cuis additive, culminating in a triple mutant Y229F/V290N/S292F‐SLAC with the highestE°′_T1Cuof 556 mV vs. SHE reported to date. Further activity assays indicate that all mutants retain oxygen reduction reaction activity, and display improved catalytic efficiencies (k_cat/K_M) relative to WT‐SLAC.