The MnCaO oxygen-evolving complex (OEC) of photosystem II catalyzes the light-driven oxidation of two substrate waters to molecular oxygen. ELDOR-detected NMR along with computational studies indicated that ammonia, a substrate analogue, binds as a terminal ligand to the Mn4A ion trans to the O5 μ oxido bridge. Results from electron spin echo envelope modulation (ESEEM) spectroscopy confirmed this and showed that ammonia hydrogen bonds to the carboxylate side chain of D1-Asp61. Here we further probe the environment of OEC with an emphasis on the proximity of exchangeable protons, comparing ammonia-bound and unbound forms. Our ESEEM and electron nuclear double resonance (ENDOR) results indicate that ammonia substitutes for the W1 terminal water ligand without significantly altering the electronic structure of the OEC.
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