Synthesis, structure and reactivity of complexes containing [M(S2)2] fragments (M=Ru, Os; (S2)=1,2-benzenedithiolate (2−))

Subsequent addition of 1,2-benzenedithiol (S₂-H₂) and nBuLi to a solution of [Ru(NO)Cl₃ · xMeOH] in THF afforded exclusively the monomeric species NBu₄[Ru^II(NO)(S₂)₂] (1). Formation of dimeric (NBu₄)₂[Ru^II(NO)(S₂)₂]₂ (2) has been confirmed when the deprotonated ligand S₂-Li₂ was added to [Ru(NO)Cl₃ · xMeOH] and allowed to stir for 30 h. The monomer 1 undergoes aerial oxidation to give (NBu₄)₂[Ru^IV(S₂)₃] (3). The reaction between RuCl₃ · xH₂O and S₂-H₂ in the presence of NaOMe, afforded the dinulear Ru^III species (NMe₄)₂[Ru^III(S₂)₂]₂ (4). A modified method for the preparation of 1 is being employed to synthesize the osmium analogue NBu₄[Os(NO)(S₂)₂] (5) effectively. The solid state structures of 1, 2 and 3 were determined by X-ray crystal structure analysis. A comparison of relevant bond distance data suggests that 1,2-benzenedithiolate acts as an “innocent” ligand.     

Carbon-13 labelling strategy for studying the ATP metabolism in individual yeast cells by micro-arrays for mass spectrometry

Isotopic labelling of cellular metabolites, used in conjunction with high-density micro-arrays for mass spectrometry enables observation of ATP metabolism in single yeast cells.     

Repetition enhancement for frequency-modulated but not unmodulated sounds: a human MEG study

Background

Decoding of frequency-modulated (FM) sounds is essential for phoneme identification. This study investigates selectivity to FM direction in the human auditory system.

Methodology/Principal Findings

Magnetoencephalography was recorded in 10 adults during a two-tone adaptation paradigm with a 200-ms interstimulus-interval. Stimuli were pairs of either same or different frequency modulation direction. To control that FM repetition effects cannot be accounted for by their on- and offset properties, we additionally assessed responses to pairs of unmodulated tones with either same or different frequency composition. For the FM sweeps, N1m event-related magnetic field components were found at 103 and 130 ms after onset of the first (S1) and second stimulus (S2), respectively. This was followed by a sustained component starting at about 200 ms after S2. The sustained response was significantly stronger for stimulation with the same compared to different FM direction. This effect was not observed for the non-modulated control stimuli.

Conclusions/Significance

Low-level processing of FM sounds was characterized by repetition enhancement to stimulus pairs with same versus different FM directions. This effect was FM-specific; it did not occur for unmodulated tones. The present findings may reflect specific interactions between frequency separation and temporal distance in the processing of consecutive FM sweeps.     

High sero-prevalence of caseous lymphadenitis identified in slaughterhouse samples as a consequence of deficiencies in sheep farm management in the state of Minas Gerais, Brazil

Background

Multiple congenital ocular anomalies (MCOA) syndrome is a hereditary congenital eye defect that was first described in Silver colored Rocky Mountain horses. The mutation causing this disease is located within a defined chromosomal interval, which also contains the gene and mutation that is associated with the Silver coat color (PMEL17, exon 11). Horses that are homozygous for the disease-causing allele have multiple defects (MCOA-phenotype), whilst the heterozygous horses predominantly have cysts of the iris, ciliary body or retina (Cyst-phenotype). It has been argued that these ocular defects are caused by a recent mutation that is restricted to horses that are related to the Rocky Mountain Horse breed. For that reason we have examined another horse breed, the Icelandic horse, which is historically quite divergent from Rocky Mountain horses.

Results

We examined 24 Icelandic horses and established that the MCOA syndrome is present in this breed. Four of these horses were categorised as having the MCOA-phenotype and were genotyped as being homozygous for the PMEL17 mutation. The most common clinical signs included megaloglobus, iris stromal hypoplasia, abnormal pectinate ligaments, iridociliary cysts occasionally extending into the peripheral retina and cataracts. The cysts and pectinate ligament abnormalities were observed in the temporal quadrant of the eyes. Fourteen horses were heterozygous for the PMEL17 mutation and were characterized as having the Cyst-phenotype with cysts and occasionally curvilinear streaks in the peripheral retina. Three additional horses were genotyped as PMEL17 heterozygotes, but in these horses we were unable to detect cysts or other forms of anomalies. One eye of a severely vision-impaired 18 month-old stallion, homozygous for the PMEL17 mutation was examined by light microscopy. Redundant duplication of non-pigmented ciliary body epithelium, sometimes forming cysts bulging into the posterior chamber and localized areas of atrophy in the peripheral retina were seen.

Conclusions

The MCOA syndrome is segregating with the PMEL17 mutation in the Icelandic Horse population. This needs to be taken into consideration in breeding decisions and highlights the fact that MCOA syndrome is present in a breed that are more ancient and not closely related to the Rocky Mountain Horse breed.     

Preparation and crystallization of a cross-linked complex of bovine adrenodoxin and adrenodoxin reductase

Bovine adrenodoxin was cross-linked to adrenodoxin reductase with 1-ethyl-3-(3-dimethyl-aminopropyl) carbodiimide. Mass spectrometry showed the reaction product to be a 1:1 complex of the two proteins with M_r = 64,790 ± 50. The cross-linked complex showed cytochrome c reductase activity and could be crystallized by hanging-drop vapor diffusion. Crystals of the adrenodoxin-adrenodoxin reductase complex are hexagonal, space group P6₁22 or P6₅22, with a = 93.26 Å and c= 612.20 Å and diffract to 2.9 Å resolution at 100 K. Assuming two cross-linked complexes per asymmetric unit yields a reasonable V_M of 2.97 ų/Da. Proteins 28:289–292, 1997. © 1997 Wiley-Liss Inc.     

Orbivirus infections in collared peccaries (Tayassu tajacu) in southeastern Brazil

We surveyed 49 free-living collared peccaries (Pecari tajacu) in Brazil for antibodies against bluetongue virus (BTV) and porcine circovirus 2 (PCV2). Antibodies against BTV were detected in 19/49 (39%) samples. All samples were negative for PCV2. The importance of antibodies to BTV in collared peccaries remains to be determined.     

Membrane topology of a metabotropic glutamate receptor

The metabotropic glutamate receptors (mGluRs) have been predicted to have a classical seven transmembrane domain structure similar to that seen for members of the G-protein-coupled receptor (GPCR) superfamily. However, the mGluRs (and other members of the family C GPCRs) show no sequence homology to the rhodopsin-like GPCRs, for which this seven transmembrane domain structure has been experimentally confirmed. Furthermore, several transmembrane domain prediction algorithms suggest that the mGluRs have a topology that is distinct from these receptors. In the present study, we set out to test whether mGluR5 has seven true transmembrane domains. Using a variety of approaches in both prokaryotic and eukaryotic systems, our data provide strong support for the proposed seven transmembrane domain model of mGluR5. We propose that this membrane topology can be extended to all members of the family C GPCRs.     

Rapid purification and crystal structure analysis of a small protein carrying two terminal affinity tags

Small peptide tags are often fused to proteins to allow their affinity purification in high-throughput structure analysis schemes. To assess the compatibility of small peptide tags with protein crystallization and to examine if the tags alter the three-dimensional structure, the N-terminus of the chicken alpha-spectrin SH3 domain was labeled with a His6 tag and the C-terminus with a StrepII tag. The resulting protein, His6-SH3-StrepII, consists of 83 amino-acid residues, 23 of which originate from the tags. His6-SH3-StrepII is readily purified by dual affinity chromatography, has very similar biophysical characteristics as the untagged protein domain and crystallizes readily from a number of sparse-matrix screen conditions. The crystal structure analysis at 2.3 A resolution proves native-like structure of His6-SH3-StrepII and shows the entire His6 tag and part of the StrepII tag to be disordered in the crystal. Obviously, the fused affinity tags did not interfere with crystallization and structure analysis and did not change the protein structure. From the extreme case of His6-SH3-StrepII, where affinity tags represent 27% of the total fusion protein mass, we extrapolate that protein constructs with N- and C-terminal peptide tags may lend themselves to biophysical and structural investigations in high-throughput regimes.     

Crystal structure analysis of an A-DNA fragment at 1.8 A resolution: d(GCCCGGGC).

Single crystals of the self-complementary octadeoxyribonucleotide d(GCCCGGGC) have been analysed by X-ray diffraction methods at a resolution of 1.8 A. The tetragonal unit cell of space group P4(3)2(1)2 has dimensions of a = 43.25 A and c = 24.61 A and contains eight strands of the oligonucleotide. The structure was refined by standard crystallographic techniques to an R factor of 17.1% using 1359 3 sigma structure factor observations. Two strands of the oligonucleotide are related by the crystallographic dyad axis to form a DNA helix in the A conformation. The d(GCCCGGGC) helix is characterized by a wide open major groove, a near perpendicular orientation of base pairs to the helix axis and an unusually small average helix twist angle of 31.3 degrees indicating a slightly underwound helix with 11.5 base pairs per turn. Extensive cross-strand stacking between guanine bases at the central cytosine-guanine step is made possible by a number of local conformational adjustments including a fully extended sugar-phosphate backbone of the central guanosine nucleotide.     

3′-5′ tRNA guanylyltransferase in bacteria

The identity of the histidine specific transfer RNA (tRNA^His) is largely determined by a unique guanosine residue at position −1. In eukaryotes and archaea, the tRNA^His guanylyltransferase (Thg1) catalyzes 3′-5′ addition of G to the 5′-terminus of tRNA^His. Here, we show that Thg1 also occurs in bacteria. We demonstrate in vitro Thg1 activity for recombinant enzymes from the two bacteria Bacillus thuringiensis and Myxococcus xanthus and provide a closer investigation of several archaeal Thg1. The reaction mechanism of prokaryotic Thg1 differs from eukaryotic enzymes, as it does not require ATP. Complementation of a yeast thg1 knockout strain with bacterial Thg1 verified in vivo activity and suggests a relaxed recognition of the discriminator base in bacteria.