@article{aaf98b1521124342a0ea89bdaa77863a,
title = "Increasing Reduction Potentials of Type 1 Copper Center and Catalytic Efficiency of Small Laccase from Streptomyces coelicolor through Secondary Coordination Sphere Mutations",
abstract = "The key to type 1 copper (T1Cu) function lies in the fine tuning of the CuII/I reduction 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 tune E°′T1Cu in 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) from Streptomyces coelicolor to match the high E°′T1Cu of 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 corresponding E°′T1Cu. While minimal and small E°′T1Cu increases are observed in Y229F- and S292F-SLAC, the V290N mutant exhibits a major E°′T1Cu increase. Moreover, the influence of these mutations on E°′T1Cu is additive, culminating in a triple mutant Y229F/V290N/S292F-SLAC with the highest E°′T1Cu of 556 mV vs. SHE reported to date. Further activity assays indicate that all mutants retain oxygen reduction reaction activity, and display improved catalytic efficiencies (kcat/KM) relative to WT-SLAC.",
keywords = "Laccase, Reduction Potential Tuning, Type 1 Copper Center",
author = "Wang, {Jing Xiang} and Vilbert, {Avery C.} and Chang Cui and Mirts, {Evan N.} and Williams, {Lucas H.} and Wantae Kim and {Jessie Zhang}, Y. and Yi Lu",
note = "This work is supported by the National Science Foundation (CHE‐2201279 for Y. L.) and National Institute for Health (R35GM148356 for Y. J. Z.). It is part of the Chemical Transformation Initiative at Pacific Northwest National Laboratory (PNNL), conducted under the Laboratory Directed Research and Development Program at PNNL, a multiprogram national laboratory operated by Battelle for the U.S. Department of Energy. We also thank the Robert A. Welch Foundation (Grant F‐0020) for support of the Lu group research program at the University of Texas at Austin. Crystallography data were acquired at the beamline 5.0.2 at the Advanced Light Source (ALS) (proposal ALS‐11565) of the Lawrence Berkeley National Laboratory, and the beamline 23‐ID‐D (proposal 81327) at the Advanced Photon Source (APS) of the Argonne National Laboratory. We would like to thank Dr. Casey van Stappen for advice in EPR spectra simulation. We would also like to thank Dr. Casey van Stappen and Mr. Yiwei Liu for their help in revising the manuscript. This work is supported by the National Science Foundation (CHE-2201279 for Y. L.) and National Institute for Health (R35GM148356 for Y. J. Z.). It is part of the Chemical Transformation Initiative at Pacific Northwest National Laboratory (PNNL), conducted under the Laboratory Directed Research and Development Program at PNNL, a multiprogram national laboratory operated by Battelle for the U.S. Department of Energy. We also thank the Robert A. Welch Foundation (Grant F-0020) for support of the Lu group research program at the University of Texas at Austin. Crystallography data were acquired at the beamline 5.0.2 at the Advanced Light Source (ALS) (proposal ALS-11565) of the Lawrence Berkeley National Laboratory, and the beamline 23-ID-D (proposal 81327) at the Advanced Photon Source (APS) of the Argonne National Laboratory. We would like to thank Dr. Casey van Stappen for advice in EPR spectra simulation. We would also like to thank Dr. Casey van Stappen and Mr. Yiwei Liu for their help in revising the manuscript.",
year = "2023",
month = dec,
day = "21",
doi = "10.1002/anie.202314019",
language = "English (US)",
volume = "62",
journal = "Angewandte Chemie - International Edition",
issn = "1433-7851",
publisher = "John Wiley & Sons, Ltd.",
number = "52",
}