Iron components in PS II particles of spinach by Mössbauer spectroscopy

Mössbauer spectrum measured for the iron components of photosystem II (PS II) particles of spinach is a superposition of 4 doublets. Quadrupole splitting and chemical shifting of doublets I–IV are characteristics of proteins with oxidized cytochrome b-559, reduced cytochrome b-559, Fe³⁺-Q complex and Fe²⁺ -Q complex respectively. After the PS II particles are treated with La³⁺, two doublets of Fe²⁺ disappear and Fe²⁺ is converted into Fe³⁺, indicating that the reduced cytochrome b-559 has been converted into the oxidized cytochrome b-559, and Fe²⁺ -Q complex into Fe³⁺ -Q complex. The Mössbauer spectrum of PS II particles treated with La³⁺ and Ca²⁺ shows that Ca²⁺ can weaken the inhibitory effect of La³⁺ in part, and a portion of the reduced cytochrome b-559 and Fe-Q complex still exist.     

Amino Acid Sequences and Distribution of High-Potential Iron–Sulfur Proteins That Donate Electrons to the Photosynthetic Reaction Center in Phototropic Proteobacteria

High-potential iron-sulfur protein (HiPIP) has recently been shown to function as a soluble mediator in photosynthetic electron transfer between the cytochrome bc1 complex and the reaction-center bacteriochlorophyll in some species of phototrophic proteobacteria, a role traditionally assigned to cytochrome c2. For those species that produce more than one high-potential electron carrier, it is unclear which protein functions in cyclic electron transfer and what characteristics determine reactivity. To establish how widespread the phenomenon of multiple electron donors might be, we have studied the electron transfer protein composition of a number of phototrophic proteobacterial species. Based upon the distribution of electron transfer proteins alone, we found that HiPIP is likely to be the electron carrier of choice in the purple sulfur bacteria in the families Chromatiaceae and Ectothiorhodospiraceae, but the majority of purple nonsulfur bacteria are likely to utilize cytochrome c2. We have identified several new species of phototrophic proteobacteria that may use HiPIP as electron donor and a few that may use cytochromes c other than c2. We have determined the amino acid sequences of 14 new HiPIPs and have compared their structures. There is a minimum of three sequence categories of HiPIP based upon major insertions and deletions which approximate the three families of phototrophic proteobacteria and each of them can be further subdivided prior to construction of a phylogenetic tree. The comparison of relationships based upon HiPIP and RNA revealed several discrepancies.     

Amino Acid Interaction with and Adsorption on Clays: FT-IR and Mössbauer Spectroscopy and X-ray Diffractometry Investigations

In the present paper, the adsorption of amino acids (Ala, Met, Gln, Cys, Asp, Lys, His) on clays (bentonite, kaolinite) was studied at different pH (3.00, 6.00, 8.00). The amino acids were dissolved in seawater, which contains the major elements. There were two main findings in this study. First, amino acids with a charged R group (Asp, Lys, His) and Cys were adsorbed on clays more than Ala, Met and Gln (uncharged R groups). However, 74% of the amino acids in the proteins of modern organisms have uncharged R groups. These results raise some questions about the role of minerals in providing a prebiotic concentration mechanism for amino acids. Several mechanisms are also discussed that could produce peptide with a greater proportion of amino acids with uncharged R groups. Second, Cys could play an important role in prebiotic chemistry besides participating in the structure of peptides/proteins. The FT-IR spectra showed that the adsorption of amino acids on the clays occurs through the amine group. However, the Cys/clay interaction occurs through the sulfhydryl and amine groups. X-ray diffractometry showed that pH affects the bentonite interlayer, and at pH 3.00 the expansion of Cys/bentonite was greater than that of the samples of ethylene glycol/bentonite saturated with Mg. The Mössbauer spectrum for the sample with absorbed Cys showed a large increase (～20%) in ferrous ions. This means that Cys was able to partially reduce iron present in bentonite. This result is similar to that which occurs with aconitase where the ferric ions are reduced to Fe 2.5.     

Study of storage iron in cultured chick embryo fibroblasts and rat glioma cells,using Mössbauer spectroscopy

⁵⁷Fe Mössbauer spectra of normal and Rous sarcoma virus-infected cultured chick embryo fibroblasts and rat glioma cells have been measured between 0.08 and 318 K. Ferritin-like iron and bacterio-ferritin-like iron have been found in these cells, in various relative amounts, indicating a close relationship between the two storage materials. The bacterio-ferritin-like iron was found to be predominantly membrane-bound. Above 260 K very wide lines were observed in the Mössbauer spectra, yielding an effective viscosity of about 1 poise in the normal chick embryo fibroblasts and about 0.5 poise in the virus-infected chick embryo fibroblasts.     

Iron-sulfur clusters of biotin synthase in vivo: a Mössbauer study

Biotin synthase, the enzyme that catalyzes the last step of the biosynthesis of biotin, contains only [2Fe-2S](2+) clusters when isolated under aerobic conditions. Previous results showed that reconstitution with an excess of FeCl(3) and Na(2)S under reducing and anaerobic conditions leads to either [4Fe-4S](2+), [4Fe-4S](+), or a mixture of [4Fe-4S](2+) and [2Fe-2S](2+) clusters. To determine whether any of these possibilities or other different cluster configuration could correspond to the physiological in vivo state, we have used (57)Fe M?ssbauer spectroscopy to investigate the clusters of biotin synthase in whole cells. The results show that, in aerobically grown cells, biotin synthase contains a mixture of [4Fe-4S](2+) and [2Fe-2S](2+) clusters. A mixed [4Fe-4S](2+):[2Fe-2S](2+) cluster form has already been observed under certain in vitro conditions, and it has been proposed that both clusters might each play a significant role in the mechanism of biotin synthase. Their presence in vivo is now another argument in favor of this mixed cluster form.     

Dynamical transition of myoglobin in a crystal: comparative studies of X-ray crystallography and Mössbauer spectroscopy

The crystallographic normal mode refinements of myoglobin at a wide range of temperature from 40 K to 300 K were carried out to study the temperature dependence of the internal atomic fluctuations. The refinement method decomposes the mean square displacement from the average position, (deltar2), into the contributions from the internal degrees of freedom and those from the external degrees of freedom. The internal displacements show linear temperature dependence as (deltar2)=alphaT+beta, throughout the temperature range measured here, and exhibit no obvious change in the slope alpha at the dynamical transition temperature (Tc=ca. 180 K). The slope alpha is practically the same as the value predicted theoretically by normal mode analysis. Such linear dependence is considered to be due to the following reason. The crystallographic Debye-Waller factor represents the static distribution caused by convolution of temperature-dependent normal mode motions and a temperature-independent set of the conformational substates. In contrast, M?ssbauer absorption spectroscopy shows a clear increase in the gradient alpha at Tc. This difference from X-ray diffraction originates from the incoherent nature of the M?ssbauer effect together with its high-energy resolution, which yields the self-correlation, and the temporal behavior of individual Fe atoms in the myoglobin crystal.     

A Comparative Study of the Spatial Distribution of Schistosomiasis in Mali in 1984–1989 and 2004–2006

Background

We investigated changes in the spatial distribution of schistosomiasis in Mali following a decade of donor-funded control and a further 12 years without control.

Methodology/Principal Findings

National pre-intervention cross-sectional schistosomiasis surveys were conducted in Mali in 1984–1989 (in communities) and again in 2004–2006 (in schools). Bayesian geostatistical models were built separately for each time period and on the datasets combined across time periods. In the former, data from one period were used to predict prevalence of schistosome infections for the other period, and in the latter, the models were used to determine whether spatial autocorrelation and covariate effects were consistent across periods. Schistosoma haematobium prevalence was 25.7% in 1984–1989 and 38.3% in 2004–2006; S. mansoni prevalence was 7.4% in 1984–1989 and 6.7% in 2004–2006 (note the models showed no significant difference in mean prevalence of either infection between time periods). Prevalence of both infections showed a focal spatial pattern and negative associations with distance from perennial waterbodies, which was consistent across time periods. Spatial models developed using 1984–1989 data were able to predict the distributions of both schistosome species in 2004–2006 (area under the receiver operating characteristic curve was typically >0.7) and vice versa.

Conclusions/Significance

A decade after the apparently successful conclusion of a donor-funded schistosomiasis control programme from 1982–1992, national prevalence of schistosomiasis had rebounded to pre-intervention levels. Clusters of schistosome infections occurred in generally the same areas accross time periods, although the precise locations varied. To achieve long-term control, it is essential to plan for sustainability of ongoing interventions, including stengthening endemic country health systems.     

Mössbauer and EPR study of iron in vacuoles from fermenting Saccharomyces cerevisiae

Vacuoles were isolated from fermenting yeast cells grown on minimal medium supplemented with 40 μM (57)Fe. Absolute concentrations of Fe, Cu, Zn, Mn, Ca, and P in isolated vacuoles were determined by ICP-MS. M?ssbauer spectra of isolated vacuoles were dominated by two spectral features: a mononuclear magnetically isolated high-spin (HS) Fe(III) species coordinated primarily by hard/ionic (mostly or exclusively oxygen) ligands and superparamagnetic Fe(III) oxyhydroxo nanoparticles. EPR spectra of isolated vacuoles exhibited a g(ave) ~ 4.3 signal typical of HS Fe(III) with E/D ~ 1/3. Chemical reduction of the HS Fe(III) species was possible, affording a M?ssbauer quadrupole doublet with parameters consistent with O/N ligation. Vacuolar spectral features were present in whole fermenting yeast cells; however, quantitative comparisons indicated that Fe leaches out of vacuoles during isolation. The in vivo vacuolar Fe concentration was estimated to be ~1.2 mM while the Fe concentration of isolated vacuoles was ~220 μM. M?ssbauer analysis of Fe(III) polyphosphate exhibited properties similar to those of vacuolar Fe. At the vacuolar pH of 5, Fe(III) polyphosphate was magnetically isolated, while at pH 7, it formed nanoparticles. This pH-dependent conversion was reversible. Fe(III) polyphosphate could also be reduced to the Fe(II) state, affording similar M?ssbauer parameters to that of reduced vacuolar Fe. These results are insufficient to identify the exact coordination environment of the Fe(III) species in vacuoles, but they suggest a complex closely related to Fe(III) polyphosphate. A model for Fe trafficking into/out of yeast vacuoles is proposed.     

Degradation of Crude Oil and PAHs in Iron–Manganese Concretions and Sediment from the Northern Baltic Sea

The ability of different sea bottom types to recover from oil contamination under oxic and anoxic conditions is poorly known in the brackish Baltic Sea. We carried out microcosm experiments to examine the capacity of iron–manganese concretions and sediment to remove petroleum compounds at 10°C. The biological degradation potential of polycyclic aromatic hydrocarbons (PAHs) was indicated by the 1000-fold increase in the copy numbers of PAH-degradation genes resulting in 2.2 × 10⁶ gene copies g^?1 DW. In the experiments 35?81% of the crude oil and 96% of PAHs disappeared through abiotic and biotic pathways, and there was little difference between the concretions and sediment. Bacterial community analysis revealed that the bacterial sequences obtained from the oil-treated concretions were affiliated to groups typical for concretions and metal-rich environments, whereas oil-treated sediment sequences were similar to those originating from sediments and oil-contaminated environments. The high frequency of concretion clones affiliated with specific taxonomic groups of α-, β-, γ-, and δ-proteobacteria may indicate the existence of new clades of bacteria within the Rhizobiales and Rhodospirillales of the α-proteobacteria, Burkholderiales—incertae sedis of the β-proteobacteria, Chromatiales of the γ-proteobacteria and the Syntrophobacteriales of the δ-proteobacteria. Our study suggests that concretions and bottom sediment maintain rich, distinct bacterial communities under oil exposure that have the potential to remove petroleum compounds in oxic and anoxic conditions.     

Iron coordination geometry in full-length, truncated, and dehydrated forms of human tyrosine hydroxylase studied by Mössbauer and X-ray absorption spectroscopy

Full-length human tyrosine hydroxylase 1 (hTH1) and a truncated enzyme lacking the 150 N-terminal amino acids were expressed in Escherichia coli and purified either with or without (6×histidine) N-terminal tags. After reconstitution with ⁵⁷Fe(II), the Mössbauer and X-ray absorption spectra of the enzymes were compared before and after dehydration by lyophilization. Before dehydration, >90% of the iron in hTH1 had Mössbauer parameters typical for high-spin Fe(II) in a six-coordinate environment [isomer shift δ(1.8–77?K)=1.26–1.24?mm s^–1 and quadrupole splitting ΔE _Q=2.68?mm s^–1]. After dehydration, the Mössbauer spectrum changed and 63% of the area could be attributed to five-coordinate high-spin Fe(II) (δ=1.07?mm s^–1 and ΔE _Q=2.89?mm s^–1 at 77?K), whereas 28% of the iron remained as six-coordinate high-spin Fe(II) (δ=1.24?mm s^–1 and ΔE _Q=2.87?mm s^–1 at 77?K). Similar changes upon dehydration were observed for truncated TH either with or without the histidine tag. After rehydration of hTH1 the spectroscopic changes were completely reversed. The X-ray absorption spectra of hTH1 in solution and in lyophilized form, and for the truncated protein in solution, corroborate the findings derived from the Mössbauer spectra. The pre-edge peak intensity of the protein in solution indicates six-coordination of the iron, while that of the dehydrated protein is typical for a five-coordinate iron center. Thus, the active-site iron can exist in different coordination states, which can be interconverted depending on the hydration state of the protein, indicating the presence or absence of a water molecule as a coordinating ligand to the iron. The present study explains the difference in iron coordination determined by X-ray crystallography, which has shown a five-coordinate iron center in rat TH, and by our recent spectroscopic study of human TH in solution, which showed a six-coordinated iron center.     

Quantum chemical calculations of spectroscopic properties of metalloproteins and model compounds: EPR and Mössbauer properties

Recently developed theoretical methods to predict EPR and M?ssbauer parameters open the way for close interactions between theorists and experimentalists to elucidate the geometric and electronic structures of metalloenzymes and model complexes and to obtain insight into their reactive properties. Spectral calculations (g-values, hyperfine couplings, zero-field splittings, isomer shifts and quadrupole splittings) are also a means to validate theoretical approaches and therefore complement the prediction of geometries, reaction energies and transition states.     

A Comparative Study on Biosurfactant Activity of Crude Oil–Degrading Bacteria and Its Correlation to Total Petroleum Hydrocarbon Degradation

In this study, 11 bacteria isolated from Tapis crude oil–contaminated sites were identified by using biochemical tests and 16S rDNA gene sequencing. Their abilities to biodegrade Tapis crude oil was determined by gas chromatography before they were further screened for biosurfactant activity by employing qualitative (blood agar hemolysis, microplate assay, drop-collapse test), semiquantitative (emulsification formation), and quantitative (surface tension measurement) methods. Four isolates, namely, Acinetobacter baumanii UKMP-12T, Pseudomonas aeruginosa UKMP-14T, Rhodococcus sp. UKMP-5T, and Rhodococcus sp. UKMP-7T, exhibited high percentages in total petroleum hydrocarbon (TPH) degradation. A strong correlation between the emulsification index (E ₂₄) and surface tension measurement (r _s = +.866) as shown by Spearman rank correlation analysis suggested that these two methods were more reliable to predict biosurfactant activity. The TPH removal was also positively correlated to the ability of bacterial isolates to reduce the surface tension of growth medium, as revealed by Pearson correlation test (r_p = +.886). In conclusion, not all the biosurfactant detection protocols employed were effective. Nevertheless, the measurement of surface tension and E ₂₄ determination provided a rather rapid, easy, reproducible, and accurate result in identifying bacteria with biosurfactant-producing ability.     

Differences in the dynamics of oxidized and reduced cytochrome c measured by Mössbauer spectroscopy

The temperature dependence of the mean square displacement of the iron atom in reduced and oxidized cytochrome c has been studied by Mössbauer spectroscopy. The flexibility of the protein, labeled by the modes coupling to the iron, is diminished upon reduction. The differences in flexibility are sufficient to explain the differences in physicochemical properties between the oxidized and the reduced forms.     

Seasonal variation in the form of iron in the liver of Spitsbergen reindeer: a Mössbauer spectroscopic study

Mössbauer spectra have been obtained from samples of the liver of Spitsbergen reindeer at different times of the year. Most of the iron is in a very similar form to that of the iron storage materials ferritin and haemosiderin. The data reflect the large differences in the amount of iron found in the liver at different times of the year and also indicate that there are only relatively small differences in the chemical and physical form of the iron found in the liver at different times of the year.     

The Role of Various Domains of the Iron–Sulfur Protein in the Assembly and Activity of the Cytochromme bc 1 Complex of Yeast Mitochondria

Assembly studies in vitro of deletion mutants of the iron–sulfur protein into the cytochromebc ₁ complex revealed that mutants localized in the extramembranous regions of the proteinwere not assembled into the complex in contrast to the efficient assembly of mutants in themembrane-spanning region. Charged amino acids located in the extramembranous 1-4 loopand the 1 helix were mutated and expressed in yeast cells lacking the gene for the iron–sulfurprotein. Mutating the charged amino acid residues H124, E125, R146, K148, and D149 aswell as V132 and W152 resulted in loss of enzymatic activity due to the loss of iron–sulfurprotein suggesting that these amino acids are required to maintain protein stability. By contrast,no loss of iron–sulfur protein accompanied the 30–50% loss of bc ₁ complex activity in mutantsof three conserved alanine residues, A86, A90, and A92, suggesting that these residues maybe involved in the proposed movement of the flexible tether of the iron–sulfur proteinduring catalysis.     

Mössbauer spectroscopic studies of deproteinised,sub-fractionated and reconstituted ferritins: the relationship between haemosiderin and ferritin

In a previous study of human haemosiderin and ferritin by a combination of Mössbauer spectroscopy and electron microscopy, it was observed that the Mössbauer spectra of haemosiderin showed a very different temperature dependence to those of ferritin. These differences were related to the superparamagnetic behaviour of small particles of a magnetic material and suggested that the magnetic anisotropy constant of the haemosiderin was considerably larger than that of the ferritin. In the present work, samples of ferritin have been examined by Mössbauer spectroscopy following partial deproteinisation, subfractionation, and reconstitution with and without phosphate, in order to investigate whether these procedures lead to changes in the magnetic anisotropy constant of the iron-containing cores. There is no evidence from the present data that changes in the protein shell, in the size of the iron-containing cores of ferritin, or in the phosphate content lead to any significant changes in the magnetic anisotropy constant, as obtained from the temperature dependence of the Mössbauer spectra. These results indicate that the different magnetic anisotropy constant observed in the case of human haemosiderin resulting from transfusional iron overload must arise from other significant differences in the composition or structure of the iron-containing cores.     

Study of storage iron in cultured chick embryo fibroblasts and rat glioma cells, using M?ssbauer spectroscopy

57Fe M?ssbauer spectra of normal and Rous sarcoma virus-infected cultured chick embryo fibroblasts and rat glioma cells have been measured between 0.08 and 318 K. Ferritin-like iron and bacterio-ferritin-like iron have been found in these cells, in various relative amounts, indicating a close relationship between the two storage materials. The bacterio-ferritin-like iron was found to be predominantly membrane-bound. Above 260 K very wide lines were observed in the M?ssbauer spectra, yielding an effective viscosity of about 1 poise in the normal chick embryo fibroblasts and about 0.5 poise in the virus-infected chick embryo fibroblasts.     

Dissolution Enhancement of a Drug Exhibiting Thermal and Acidic Decomposition Characteristics by Fusion Processing: A Comparative Study of Hot Melt Extrusion and KinetiSol® Dispersing

In this study, hot melt extrusion (HME) and KinetiSol® Dispersing (KSD) were utilized to prepare dissolution-enhanced solid dispersions of Roche Research Compound A (ROA), a BCS class II drug. Preformulation characterization studies showed that ROA was chemically unstable at elevated temperatures and acidic pH values. Eudragit® L100-55 and AQOAT® LF (HPMCAS) were evaluated as carrier polymers. Dispersions were characterized for ROA recovery, crystallinity, homogeneity, and non-sink dissolution. Eudragit® L100-55 dispersions prepared by HME required the use of micronized ROA and reduced residence times in order to become substantially amorphous. Compositions containing HPMCAS were also prepared by HME, but an amorphous dispersion could not be obtained. All HME compositions contained ROA-related impurities. KSD was investigated as a method to reduce the decomposition of ROA while rendering compositions amorphous. Substantially amorphous, plasticizer free compositions were processed successfully by KSD with significantly higher ROA recovery values and amorphous character than those achieved by HME. A near-infrared chemical imaging analysis was conducted on the solid dispersions as a measure of homogeneity. A statistical analysis showed similar levels of homogeneity in compositions containing Eudragit® L100-55, while differences were observed in those containing HMPCAS. Non-sink dissolution analysis of all compositions showed rapid supersaturation after pH adjustment to approximately two to three times the equilibrium solubility of ROA, which was maintained for at least 24 h. The results of the study demonstrated that KSD is an effective method of forming dissolution-enhanced amorphous solid solutions in cases where HME is not a feasible technique.