Direct Comparison of CuB and CuSiR Sites in a Single Designed Protein Scaffold Reveals a Central Role for Tyrosine in Oxygen and Sulfite Reduction.

Authors

Aaron P Ledray, Barshali Ghosh, Hirbod Heidari, Yi Lu

Published in

Journal of the American Chemical Society. Volume 148. Issue 32. Pages 34094-34101. Aug 19, 2026.

Abstract

Both heme-copper oxidase (HCO) and sulfite reductase type A (SiR-A) enzymes rely on a copper center next to heme to perform their biological reactions, but a question remains as to why the CuB center in HCO promotes the oxygen reduction reaction (ORR) to water that is essential for generating a proton motive force to synthesize ATP in aerobic life, while the CuSiR in SiR-A is efficient at the sulfite reduction reaction (SRR) to sulfide, generating a proton motive force in anaerobic bacteria. To answer this question, we report herein the design of either a CuB or a CuSiR site in a single protein, soybean ascorbate peroxidase (APX), as a scaffold to mimic those in HCO and SiR-A, respectively. By eliminating confounding factors from different native scaffolds, we isolate the intrinsic roles of copper coordination environments and a conserved Tyr in catalyzing fundamentally different reactions, offering unprecedented clarity on how these centers govern these reactions. Our results reveal that both CuB and CuSiR sites can support ORR, but only CuSiR promotes SRR, underscoring the greater challenge of 6e- /7H+ transfer versus 4e- /4H+. Surprisingly, introducing a Tyr near the CuB site not only enhances ORR but also confers robust SRR activity, surpassing even CuSiR under identical conditions. This finding changed the view that copper coordination alone dictates catalytic specificity and highlights the dominant role of Tyr-Cu synergy in enabling multielectron redox chemistry. These insights provide a new framework for understanding evolutionary design principles and for creating bioinspired catalysts for selective O2 and SO32- reductions.

PMID:
42619153
Bibliographic data and abstract were imported from PubMed on 20 Aug 2026.

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