Location of Chlorophyll_z in Photosystem II

Saturation-recovery and progressive microwave power saturation EPR spectroscopies have been used to probe the location of the chlorophyll_z⁺ (Chl_z⁺) radical species in Mn-depleted photosystem II (PSII). The spin-lattice relaxation transients of Chl_z⁺ were non-single-exponential due to a dipole-dipole interaction with one of the other paramagnetic centers in PSII. Measurements on CN^--treated, Mn-depleted PSII membrane samples, in which the non-heme Fe(II) is converted into its low-spin, diamagnetic form, confirmed that the non-heme Fe(II) caused the dipolar relaxation enhancement of Chl_z⁺. The saturation-recovery EPR data were fit to a dipolar model [Hirsh, D. J., Beck, W. F., Innes, J. B., & Brudvig, G. W. (1992) Biochemistry 31, 532] which takes into account the isotropic (scalar) and orientationdependent (dipolar) contributions to the spin-lattice relaxation of the radical. The temperature dependence of the dipolar rate constants of Chl_z⁺was identical to the temperature dependencies recently observed for the stable tyrosine radical, Y_d^•, and the special pair bacteriochlorophyll radical, (BChla)₂⁺, in PSII and in reaction centers from Rhodobacter sphaeroides, respectively. Because the non-heme Fe(II) is known to cause a dipolar relaxation enhancement of the radicals in both of the latter cases, this result provides further evidence that the non-heme Fe(II) causes the dipolar relaxation enhancement of Chl_z⁺ and, moreover, demonstrates that the magnetic properties of the non-heme Fe(II) in PSII and in reaction centers from Rhodobacter sphaeroides are very similar. By using the known Fe(II)-(BChla)₂⁺ distance for calibration, we estimate the Fe(II)-Chl_z⁺ distance to be 39.5 ± 2.5 Å. The theory of dipolar relaxation enhancement of a free radical caused by exogenous Dy³⁺ complexes [Innes, J. B., & Brudvig, G. W. (1989) Biochemistry 28, 1116] has also been applied to determine the location of Chl_z⁺ relative to the PSII protein surfaces. Chl_z⁺ was found to be located at approximately equal distances from both the luminal and stromal protein surfaces in extrinsic polypeptide-depleted PSII membranes. These results provide the first direct evidence against the assignment of Chlz to a Chi monomer analogous to the “voyeur” BChl in the bacterial reaction center and point to histidines-118 in the DI and D2 proteins as potential ligands of Chl_z.