The dynamics of the non-heme iron in bacterial reaction centers from Rhodobacter sphaeroides.

We investigate the dynamical properties of the non-heme iron (NHFe) in His-tagged photosynthetic bacterial reaction centers (RCs) isolated from Rhodobacter (Rb.) sphaeroides. Mössbauer spectroscopy and nuclear inelastic scattering of synchrotron radiation (NIS) were applied to monitor the arrangement and flexibility of the NHFe binding site. In His-tagged RCs, NHFe was stabilized only in a high spin ferrous state. Its hyperfine parameters (IS=1.06±0.01mm/s and QS=2.12±0.01mm/s), and Debye temperature (θ(D0)~167K) are comparable to those detected for the high spin state of NHFe in non-His-tagged RCs. For the first time, pure vibrational modes characteristic of NHFe in a high spin ferrous state are revealed. The vibrational density of states (DOS) shows some maxima between 22 and 33meV, 33 and 42meV, and 53 and 60meV and a very sharp one at 44.5meV. In addition, we observe a large contribution of vibrational modes at low energies. This iron atom is directly connected to the protein matrix via all its ligands, and it is therefore extremely sensitive to the collective motions of the RC protein core. A comparison of the DOS spectra of His-tagged and non-His-tagged RCs from Rb. sphaeroides shows that in the latter case the spectrum was overlapped by the vibrations of the heme iron of residual cytochrome c(2), and a low spin state of NHFe in addition to its high spin one. This enabled us to pin-point vibrations characteristic for the low spin state of NHFe.

Coupling of collective motions of the protein matrix to vibrations of the non-heme iron in bacterial photosynthetic reaction centers.

Non-heme iron is a conservative component of type II photosynthetic reaction centers of unknown function. We found that in the reaction center from Rba. sphaeroides it exists in two forms, high and low spin ferrous states, whereas in Rsp. rubrum mostly in a low spin state, in line with our earlier finding of its low spin state in the algal photosystem II reaction center (Burda et al., 2003). The temperature dependence of the non-heme iron displacement studied by Mössbauer spectroscopy shows that the surrounding of the high spin iron is more flexible (Debye temperature ~165K) than that of the low spin atom (~207K). Nuclear inelastic scattering measurements of the collective motions in the Rba. sphaeroides reaction center show that the density of vibrational states, originating from non-heme iron, has well-separated modes between lower (4-17meV) and higher (17-25meV) energies while in the one from Rsp. rubrum its distribution is more uniform with only little contribution of low energy (~6meV) vibrations. It is the first experimental evidence that the fluctuations of the protein matrix in type II reaction center are correlated to the spin state of non-heme iron. We propose a simple mechanism in which the spin state of non-heme iron directly determines the strength of coupling between the two quinone acceptors (Q(A) and Q(B)) and fast collective motions of protein matrix that play a crucial role in activation and regulation of the electron and proton transfer between these two quinones. We suggest that hydrogen bond network on the acceptor side of reaction center is responsible for stabilization of non-heme iron in different spin states.

Interaction of chlorpromazine, fluphenazine and trifluoperazine with ocular and synthetic melanin in vitro.

The aim of this study was to examine in vitro the binding capacity of three phenothiazine derivatives--chlorpromazine, fluphenazine and trifluoperazine--causing adverse effects in the eye structures, to natural melanin isolated from pig eyes as well as to synthetic DOPA-melanin used as a model polymer. The amount of drug bound to melanin was determined by UV spectrophotometry. The analysis of results for the kinetics of drug-melanin complex formation showed that the amount of drug bound to melanin increases with increasing initial drug concentration and longer incubation time, attaining an equilibrium state after about 24 h. Binding parameters, i.e. the number of binding sites (n) and association constants (K), were determined on the basis of Scatchard plots. For neuroleptic-ocular melanin and neuroleptic-DOPA-melanin complexes two classes of independent binding sites were found, with association constants K1 approximately 10(4) and K2 approximately 10(2) M (-1) for chlorpromazine and fluphenazine complexes, and K1 approximately 10(5) and K2 approximately 10(3) M(-1) for trifluoperazine complexes. The numbers of strong (n1) and weak (n2) binding sites indicate lower affinity of the drugs examined to ocular melanin compared with DOPA-melanin. The ability of chlorpromazine, fluphenazine and trifluoperazine to interact with melanin, especially the ocular melanin, in vitro is discussed in relation to the ocular toxicity of these drugs in vivo.

The influence of selected metals and halogens on the electronic system of benzoic acid.

The influence of halogens and metals on the electronic system of the aromatic ring in lithium, sodium and potassium complexes with p-halogenobenzoic acids has been investigated by means of (13)C and (1)H NMR, IR and Raman spectroscopy and semi-empirical calculations. It has been shown that ionic potentials and electronegativities of halogens and metals are the main factors responsible for perturbations of the electronic charge distribution in the ring.