Construction of carbon chains on ruthenium and osmium carbonyl clusters

We have used the elimination of AuX(PR₃) (X = halide, R = Ph, tol) that occurs in reactions of alkynylgold(I)-phosphine complexes with M₃(μ-H)₃ (μ₃-CBr) (CO)₉ (M = Ru, Os) to prepare the complexes M₃(μ-H)₃(μ₃-CCCR)(CO)₉ [M = Ru, R = Ph 2, CCSiMe₃3, Fc 4, CCFc 6-Ru, CC[Ru(PPh₃)₂Cp] 8; M = Os, R = CCFc 6-Os, CCCCFc 7], Fc′{(μ₃-CCC)Ru₃(μ-H)₃(CO)₉}₂5, and bis-cluster-capped carbon chain complexes {M₃(μ-H)₃(CO)₉}₂{μ₃:μ₃-C(CC)_nC} (M = Ru, n = 2 9, 3 10-Ru; M = Os, n = 3 10-Os) and {(L)(OC)₈(μ-H)₃M₃}C(CC)_nC{Co₃(μ-dppm)(CO)₇} (n = 1, M = Ru, L = CO 11, PPh₃12-Ru/P; n = 2, L = CO 12-Ru, PPh₃13; M = Os, L = CO 12-Os) in good to excellent yields. X-ray structural determinations of 2-5, 6-Ru, 6-Os, 7, 9, 11, 12-Ru, 12-Os and 12-Ru/P are reported.     

Mass tag-assisted identification of naturally processed HLA class II-presented meningococcal peptides recognized by CD4+ T lymphocytes

The meningococcal class I outer membrane protein porin A plays an important role in the development of T cell-dependent protective immunity against meningococcal serogroup B infection and is therefore a major component of candidate meningococcal vaccines. T cell epitopes from porin A are poorly characterized because of weak in vitro memory T cell responses against purified Ag and strain variation. We applied a novel strategy to identify relevant naturally processed and MHC class II-presented porin A epitopes, based on stable isotope labeling of Ag. Human immature HLA-DR1-positive dendritic cells were used for optimal uptake and MHC class II processing of (14)N- and (15)N-labeled isoforms of the neisserial porin A serosubtype P1.5-2,10 in bacterial outer membrane vesicles. HLA-DR1 bound peptides, obtained after 48 h of Ag processing, contained typical spectral doublets in mass spectrometry that could easily be assigned to four porin A regions, expressed at diverging densities ( approximately 30-4000 copies/per cell). Epitopes from two of these regions are recognized by HLA-DR1-restricted CD4(+) T cell lines and are conserved among different serosubtypes of meningococcal porin A. This mass tag-assisted approach provides a useful methodology for rapid identification of MHC class II presented bacterial CD4(+) T cell epitopes relevant for vaccine development.     

Identification of formaldehyde-induced modifications in proteins: reactions with model peptides

Formaldehyde is a well known cross-linking agent that can inactivate, stabilize, or immobilize proteins. The purpose of this study was to map the chemical modifications occurring on each natural amino acid residue caused by formaldehyde. Therefore, model peptides were treated with excess formaldehyde, and the reaction products were analyzed by liquid chromatography-mass spectrometry. Formaldehyde was shown to react with the amino group of the N-terminal amino acid residue and the side-chains of arginine, cysteine, histidine, and lysine residues. Depending on the peptide sequence, methylol groups, Schiff-bases, and methylene bridges were formed. To study intermolecular cross-linking in more detail, cyanoborohydride or glycine was added to the reaction solution. The use of cyanoborohydride could easily distinguish between peptides containing a Schiff-base or a methylene bridge. Formaldehyde and glycine formed a Schiff-base adduct, which was rapidly attached to primary N-terminal amino groups, arginine and tyrosine residues, and, to a lesser degree, asparagine, glutamine, histidine, and tryptophan residues. Unexpected modifications were found in peptides containing a free N-terminal amino group or an arginine residue. Formaldehyde-glycine adducts reacted with the N terminus by means of two steps: the N terminus formed an imidazolidinone, and then the glycine was attached via a methylene bridge. Two covalent modifications occurred on an arginine-containing peptide: (i) the attachment of one glycine molecule to the arginine residue via two methylene bridges, and (ii) the coupling of two glycine molecules via four methylene bridges. Remarkably, formaldehyde did not generate intermolecular cross-links between two primary amino groups. In conclusion, the use of model peptides enabled us to determine the reactivity of each particular cross-link reaction as a function of the reaction conditions and to identify new reaction products after incubation with formaldehyde.     

Mutations in the VLGR1 gene implicate G-protein signaling in the pathogenesis of Usher syndrome type II

Usher syndrome type II (USH2) is a genetically heterogeneous autosomal recessive disorder with at least three genetic subtypes (USH2A, USH2B, and USH2C) and is classified phenotypically as congenital hearing loss and progressive retinitis pigmentosa. The VLGR1 (MASS1) gene in the 5q14.3-q21.1 USH2C locus was considered a likely candidate on the basis of its protein motif structure and expressed-sequence-tag representation from both cochlear and retinal subtracted libraries. Denaturing high-performance liquid chromatography and direct sequencing of polymerase-chain-reaction products amplified from 10 genetically independent patients with USH2C and 156 other patients with USH2 identified four isoform-specific VLGR1 mutations (Q2301X, I2906FS, M2931FS, and T6244X) from three families with USH2C, as well as two sporadic cases. All patients with VLGR1 mutations are female, a significant deviation from random expectations. The ligand(s) for the VLGR1 protein is unknown, but on the basis of its potential extracellular and intracellular protein-protein interaction domains and its wide mRNA expression profile, it is probable that VLGR1 serves diverse cellular and signaling processes. VLGR1 mutations have been previously identified in both humans and mice and are associated with a reflex-seizure phenotype in both species. The identification of additional VLGR1 mutations to test whether a phenotype/genotype correlation exists, akin to that shown for other Usher syndrome disease genes, is warranted.     

Site-specific labeling of ‘second generation’ annexin V with 99mTc(CO)3 for improved imaging of apoptosis in vivo

In this study ‘second generation’ AnxV was specifically labeled with ^99mTc in three different ways outside the binding region of the protein to obtain an improved target-to-background activity ratio. The compounds were tested in vitro and in vivo in normal mice and in a model of hepatic apoptosis (anti-Fas mAb). The apoptosis binding was most prominent for the HIS-tagged ‘second generation’ AnxV labeled with ^99mTc(CO)₃ in comparison to ^99mTc-HYNIC-cys-AnxV and ^99mTc(CO)₃-DTPA-cys-AnxV.     

Potential impact of grey squirrels Sciurus carolinensis on woodland bird populations in England

Several woodland bird species have declined markedly in abundance in England over the past 40 years, whilst the grey squirrel Sciurus carolinensis, a non-native nest predator, has increased. Given the timing, there has been concern that grey squirrels have driven these declines, although there is little data to support this view. Using novel analytical methods and extensive national bird and grey squirrel monitoring data, we examine whether there is evidence that woodland bird populations in England have been depressed by grey squirrels and whether there is a relationship between nest failure and squirrel numbers. Our results indicate that grey squirrels are very unlikely to have driven observed declines of woodland birds in recent years, although the number of associations, positive as well as negative, between grey squirrels and woodland birds is greater than expected by chance. For this reason, we cannot exclude the possibility that the populations of a small number of bird species, principally increasing species, have been depressed to some degree at sites where a greater number of grey squirrels were present. Of these species, perhaps the most convincing evidence is for Common Blackbird Turdus merula and Eurasian Collared Dove Streptopelia decaocto where nest record data also identified a positive relationship between nest failure at the egg stage and squirrel abundance.     

The relationship of accentuated lines in enamel to weaning stress in juvenile baboons (Papio hamadryas anubis)

Weaning conflict may represent an evolutionary conflict of interest between parent and offspring, an honest signal of need on the part of a weanling, or both. Accentuated lines visible in histological sections of teeth are indicators of stress during enamel formation and have been hypothesised to form in baboon teeth during weaning. We analysed growth increments in 5 tooth sections from 2 Ugandan baboons (Papio hamadryas anubis), using polarised light microscopy, to determine when stresses occurred during the weaning process. Dietary transitions were reconstructed using normalised strontium intensities (Sr/Ca) in enamel. Accentuated lines were cross-matched between teeth from the same animal and plotted by month. The highest frequency of stress was experienced at around 6 months in 1 baboon, coinciding with an inferred reduction in suckling frequency, and at 11 months in another, coinciding with the inferred cessation of suckling. Because accentuated lines appear to indicate weaning stress at dietary transitions, weaning conflict may represent an honest signal of need on the part of the weanling.     

Effect of MELANOTAN, [Nle(4), D-Phe(7)]-alpha-MSH, on melanin synthesis in humans with MC1R variant alleles

MELANOTAN (NDP-MSH) binds the MC1 receptor to significantly increase the eumelanin content of human skin cells. In this study of 77 Caucasian individuals, we investigated the effects of MELANOTAN in individuals with variant MC1R genotypes, as it has been suggested through in vitro studies that variant alleles decrease MELANOTAN binding efficacy, which would subsequently affect the synthesis of melanin. Administration of MELANOTAN produced a significant (p<0.001) increase in melanin density in treated, compared to placebo, individuals. Importantly, MELANOTAN increased the melanin density to a greater extent in individuals carrying the variant alleles Val60Leu, Asp84Glu, Val92Met, Arg142His, Arg151Cys, and Arg160Trp than in individuals with no variant alleles. This study demonstrates that MELANOTAN effectively increases the melanin content of skin in those individuals with MC1R variant alleles and therefore, those most in need of photoprotection.     

Strigol: biogenesis and physiological activity

The role played by molecules of the strigolactone family in stimulating the germination of seeds of parasitic weeds of the genera Striga, Orobanche and Alectra has never been clearly elucidated. The biogenesis of these unusual terpenoid lactones, originally identified in minute quantities in the root exudates of a small number of host plants and two or three "false hosts", also remains obscure. These lactones, as the chemical signals which initiate the life cycle of Striga, are consequently at the forefront of the Striga research effort. This paper reviews recent key discoveries relating to the biosynthesis and mode of action of strigolactones, and summarises the evidence suggesting that these molecules may be far more widely distributed and have a greater physiological significance than has hitherto been appreciated.     

Sources of Campylobacter spp. colonizing housed broiler flocks during rearing

The study aimed to identify sources of campylobacter in 10 housed broiler flocks from three United Kingdom poultry companies. Samples from (i) the breeder flocks, which supplied the broilers, (ii) cleaned and disinfected houses prior to chick placement, (iii) the chickens, and (iv) the environments inside and outside the broiler houses during rearing were examined. Samples were collected at frequent intervals and examined for Campylobacter spp. Characterization of the isolates using multilocus sequence typing (MLST), serotyping, phage typing, and flaA restriction fragment length polymorphism typing was performed. Seven flocks became colonized during the growing period. Campylobacter spp. were detected in the environment surrounding the broiler house, prior to as well as during flock colonization, for six of these flocks. On two occasions, isolates detected in a puddle just prior to the birds being placed were indistinguishable from those colonizing the birds. Once flocks were colonized, indistinguishable strains of campylobacter were found in the feed and water and in the air of the broiler house. Campylobacter spp. were also detected in the air up to 30 m downstream of the broiler house, which raises the issue of the role of airborne transmission in the spread of campylobacter. At any time during rearing, broiler flocks were colonized by only one or two types determined by MLST but these changed, with some strains superseding others. In conclusion, the study provided strong evidence for the environment as a source of campylobacters colonizing housed broiler flocks. It also demonstrated colonization by successive campylobacter types determined by MLST during the life of a flock.