THE STATE OF MANGANESE IN THE PHOTOSYNTHETIC APPARATUS - NITROGEN LIGATION TO MANGANESE IN THE PHOTOSYNTHETIC OXYGEN-EVOLVING COMPLEX - CONTINUOUS-WAVE AND PULSED EPR STUDIES OF PHOTOSYSTEM-II PARTICLES CONTAINING N-14 OR N-15

The possibility of nitrogen ligation to the Mn in the oxygen-evolving complex from photosystem II was investigated with electron paramagnetic resonance (EPR) and electron spin echo envelope modulation (ESEEM) spectroscopies using N-14- and N-15-labeled preparations. Oxygen-evolving preparations were isolated from a thermophilic cyanobacterium, Synechococcus sp., grown on a medium containing either (NO3-)-N-15 as the sole source of nitrogen. The substructure on the "multiline" EPR signal, which arises from Mn in the S2 state of the enzyme, was measured with continous-wave EPR. No changes were detected in the substructure peak positions upon substitution of N-15 for N-14, indicating that this substructure is not due to superhyperfine coupling from nitrogen ligands. To detect potential nitrogen ligands with superhyperfine couplings of lesser magnitude than could be observed with conventional EPR methods, electron spin-echo envelope modulation experiments were also performed on the multiline EPR signal. The Fourier transform of the light-minus-dark time domain ESEEM data shows a peak at 4.8 MHz in N-14 samples which is absent upon substitution with N-15. This gives unambiguous evidence for weak hyperfine coupling of nitrogen to the Mn of the oxygen-evolving complex. Possible origins of this nitrogen interaction are discussed.