Comparative FTIR analysis of the microenvironment of QA-• in cyanide-treated, high pH-treated and iron-depleted photosystem II membrane fragments

FTIR difference spectra due to light-induced Q(A)(-.) formation were measured in control PS II membrane fragments and in samples where the magnetic interaction with the non-heme iron center in its high-spin Fe2+ state is eliminated by three different methods, i.e., extraction of the non-heme iron center, treatment with cyanide, and incubation at high pH (pH = 11). The results obtained reveal that (i) the most pronounced band at 1479 cm(-1) reflecting the Cchemical anionO stretching mode of Q(A)(-.) in the semiquinone anion radical remains invariant to "iron depletion" while it shifts by 4 and 2 cm(-1) to lower frequencies upon cyanide and high pH treatments, respectively, (ii) peaks observed in the 2600-3000 cm(-1) region which arise from Fermi resonance of harmonics and combinations of the imidazole ring modes with the hydrogen-bonding NH stretching vibrations are not affected upon iron depletion but are lost in cyanide and high pH-treated samples, and (iii) all three treatments give rise to some similar changes in the amide I bands of the protein backbones and in imidazole ring modes of the coupled histidine. These results show that the hydrogen-bonding interaction of Q(A)(-.) is virtually unaffected upon non-heme iron depletion; in particular, the strong hydrogen bond between Q(A) and a histidine side chain (most likely His 215 of the D2 subunit) is not changed. In marked contrast, drastic changes take place in the hydrogen bonding between Q(A) and His upon CN- and high pH treatments. The straightforward interpretation is that the hydrogen bond is lost upon these treatments. Despite the striking difference in the effect of hydrogen-bonding interaction, all three treatments lead to similar structural pertubations on the protein conformational changes due to Q(A)(-.) formation and ring vibrations of the coupled histidine side chain. On the basis of the data presented in the study, it is inferred that, concerning the hydrogen bond interaction, the microenvironment of Q(A)(-.) is close to the native state when a suitable iron depletion is performed. Accordingly, the previously reported conclusion on the hydrogen-bonding pattern of Q(A)(-.) in PS II [MacMillan, F., Lendzian, F., Renger, G., and Lubitz, W. (1995) Biochemistry 34, 8144-8156] studied by using iron-depleted preparations most likely reflects the situation in an intact PS II.