| Abstract
| - The non heme iron environment of photosystem II is studied by light-induced infraredspectroscopy. A conclusion of previous work [Hienerwadel, R., and Berthomieu, C. (1995) Biochemistry34, 16288−16297] is that bicarbonate is a bidendate ligand of the reduced iron and a monodentate ligandin the Fe3+ state. In this work, the effects of bicarbonate replacement with lactate, glycolate, and glyoxylate,and of o-phenanthroline binding are investigated to determine the specific interactions of bicarbonatewith the protein. Fe2+/Fe3+ FTIR spectra recorded with 12C- and 13C1-labeled lactate indicate that lactatedisplaces bicarbonate by direct binding to the iron through one carboxylate oxygen and the hydroxylgroup in both the Fe2+ and Fe3+ states. This different binding mode with respect to bicarbonate couldexplain the lower midpoint of the iron couple observed in the presence of this anion [Deligiannakis, Y.,Petrouleas, V., and Diner, B. A. (1994) Biochim. Biophys. Acta 1188, 260−270]. In agreement with the−60 mV/pH unit dependence of the iron midpoint potential in the presence of bicarbonate, the protonrelease upon iron oxidation by photosystem II is directly measured to 0.95 ± 0.05 by the comparison ofinfrared signals of phosphate buffer and ferrocyanide modes. This accurate method may be applied to thestudy of other redox reactions in proteins. The pH dependence of the iron couple is proposed to reflectthe deprotonation of D1His215, a putative iron ligand located at the QB pocket, since the signal at 1094cm-1 assigned to the ν(C−N) mode of a histidinate ligand in the Fe3+ state is not observed in the presenceof o-phenanthroline. Specific regulation of the pKa of D1His215 by bicarbonate is inferred from the absenceof the band at 1094 cm-1 in Fe2+/Fe3+ spectra recorded with glycolate, glyoxylate, or lactate. A broadpositive continuum, maximum at ≈2550 cm-1, observed in the presence of bicarbonate, but absent witho-phenanthroline or lactate, glycolate, and glyoxylate, indicates a hydrogen bond network from the nonheme iron toward the QB pocket involving bicarbonate and His D1−215. Proton release of about 1,measured upon iron oxidation at pH 6 with the latter anions, points to a proton release mechanism differentfrom that involved in the presence of bicarbonate.
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