Amino Acids

Previous experimental studies have highlighted that citrulline (CIT) could be a promising pharmaconutrient. However, its pharmacokinetic characteristics and tolerance to loading have not been studied to date. The objective was to characterise the plasma kinetics of CIT in a multiple-dosing study design and to assess the effect of CIT intake on the concentrations of other plasma amino acids (AA). The effects of CIT loading on anabolic hormones were also determined. Eight fasting healthy males underwent four separate oral loading tests (2, 5, 10 or 15 g CIT) in random order. Blood was drawn ten times over an 8 h period for measurement of plasma AA, insulin and growth hormone (Gh). Urine samples were collected before CIT administration and over the next 24 h. None of the subjects experienced side effects whatever the CIT dose. Concerning AA, only CIT, ornithine (ORN) and arginine (ARG) plasma concentrations were affected (maximum concentration 146 (sem 8) to 303 (sem 11) μmol/l (ARG) a...

For an inconclusive study of unimolecular cleavage of cyclopropanes, see: A. 6. Chmurny and D. J. Cram, J. Am. Chem. SOC., 95,4237 (1973), and previous papers in this series. (27) The first examples of reliance on configurationally defined cyclopropanes with intramolecular displacement as a strategy in stereospecific synthesis were provided by G. Stork and associates, see: G. Stork and M. Marx, J. Am. Chem. Soc., 91, 237 1 (1969); G. Stork and M. Gregson, ibid., 91,2373Gregson, ibid., 91, (1969. It is interesting to note that those ring openings which involved acid catalysis occurred strictly in the fused mode. (28) J. Dynak. Ph.D. Thesis, University of Pittsburgh, 1975. (29) D. Landine and R. Rolla, Synthesis, 389 (1976). (30) (a) S. Tripett and D. M. Walker, J. Chem. Soc., 1266 (1961); (b) M. Regitz, Chem. Ber., 99, 3128 (1966); (c) these conditions were provided to us by Professor 6. M. Trost. (31) E. N. Marvel1 and T. Li, Synthesis, 457 (1973). (32) The 250-MHz spectra of lactam alcohols 25 and 26 and of trachelanthamidine and isoretronecanol are provided in the microfilm edition (consult masthead page for ordering information). (33) Combustion analyses were performed by Galbraith Laboratories, Knoxville Tenn. Infrared spectra were measured on a Perkin-Elmer 137 lnfracord spectrophotometer. NMR spectra were obtained at 60 MHz on Varian A-60D cf T-60 systems and at 250 MHz as indicated, with tetramethylsilane as an internal standard. Data are reported in 6 (ppm) from the Me&i signal. Melting points are uncorrected.

The synthesis of a limited series of non-racemic atropisomeric 1-(2-(4methyl-2-thioxothiazol-3(2H)-yl)phenyl)-3-(hetero)aryl-(thio)ureas is described. Using NMR titration experiments monitoring the shift of the two NH of the (thio)urea and the C-5 hydrogen of the heterocycle, the binding constants for some optically pure (thio)ureas with the enantiomers of N-protected amino acid tetrabutylammonium salts were determined in CD 3 CN. The obtained enantioselectivities were modest. Contrary to what was expected on the basis of the NH acidity in thiourea versus urea group, the association constants were smaller with the thiourea than with the corresponding urea. X-ray data, DFT calculations, and NMR provided the explanation of that unexpected behavior: the urea presents a pre-organized (Z,Z) conformation suitable for a double hydrogen bond with the carboxylate anion, the thiourea presents a (Z,E) conformation, which must be reorganized in a constrained (Z,Z) conformation in the complex. An intramolecular hydrogen bond between one NH and the thiocarbonyl group of the heterocycle, which is present in the thiourea and absent in the urea, might also contribute to the smaller K ass for the thiourea. The possible implication of these observations in the field of bifunctional organocatalysis is briefly discussed.

Circadian rhythm abnormalities have been described in various psychiatric disorders, but they have not received much attention in studies of schizophrenia and schizophreniform psychosis. The present study used the cosine model to determine the circadian patterns of amino acids, dopamine, and prolactin concentrations, which were analyzed over a 24-hour period in serum of healthy subjects, drug-free schizophrenic patients, and neuroleptic-treated schizophrenic patients. The mesor (the daily mean) of phenylalanine was lower in drug-free schizophrenic women than in healthy women. The mesors of the ratio of phenylalanine or tyrosine to competing amino acids were similar in healthy subjects and patients. The ratio of phenylalanine/competing amino acids showed a phase advance (i.e., earlier onset of the time of highest concentration) in drugfree patients compared with healthy subjects. Schizophrenic patients displayed a higher dopamine mesor than healthy subjects. Female drug-free schizophrenic patients had lower prolactin mesors and lower amplitudes (i.e., half of the total predictable change in rhythm) than healthy women. Compared with healthy subjects, schizophrenic patients showed a phase advance of circadian prolactin concentrations. Neuroleptics raised the prolactin mesor and amplitudes but did not elicit any phase change in amino acids, dopamine, or prolactin. These data confirm the indirect pharmacologic evidence of increased dopaminergic activity in schizophrenic patients that relates to dopamine's precursors and to the neuroendocrine regulation of prolactin.

High pressure homogenization (HPH) was tested for inducing autolysis in a commercial strain of Saccharomyces bayanus for winemaking. The effects on cell viability, the release of soluble proteins, glucidic colloids and amino acids in wine-like medium and the volatile composition of the autolysates were investigated after processing, in comparison with thermolysis. HPH seemed a promising technique for inducing autolysis of wine yeasts. One pass at 150MPa was the best operating conditions. Soluble colloids, proteins and free amino acids were similar after HPH and thermolysis, but the former gave a more interesting volatile composition after processing, with higher concentrations of ethyl esters (fruity odors) and lower fatty acids (potential off-flavors). This might allow different winemaking applications for HPH, such as the production of yeast derivatives for wine ageing. In the conditions tested, HPH did not allow the complete inactivation of yeast cells; the treatment shall be optimized before winemaking use.

In this report, we have investigated the contribution of primary sequence to the carbohydrate requirement for intracellular transport of two closely related glycoproteins, the G proteins of the San Juan and Orsay strains of vesicular stomatitis virus. We used site-directed mutagenesis of the coding sequence to eliminate the two consensus sites for glycosylation in the Orsay G protein. Whereas the nonglycosylated San Juan G protein required at least one of its two asparagine-linked oligosaccharides for transport to the plasma membrane at 37 degrees C, a fraction of the Orsay G protein was transported without carbohydrate. Of the 10 amino acid differences between these two proteins, residue 172 (tyrosine in San Juan, aspartic acid in Orsay) played the major role in determining the stringency for the carbohydrate requirement. The rates at which the glycosylated and nonglycosylated Orsay G proteins were transported to the cell surface were the same, although a smaller fraction of the no...

Genetic mutations affecting mitochondrial fission and fusion proteins cause human neurological disorders, but are assumed to be well tolerated in yeast. The conserved mitochondrial fission protein Dnm1/Drp1 is required for normal mitochondrial division, but also promotes cell death in mammals and yeast. Fis1, an outer mitochondrial membrane-anchored receptor for Dnm1/Drp1, also can promote cell death in mammals, but appears to have prosurvival activity in yeast. Here we report that deletion of the FIS1 gene in yeast consistently results in acquisition of a secondary mutation that confers sensitivity to cell death. In several independently derived FIS1 knockouts, tiling arrays and genomic sequencing identified the secondary mutation as a premature termination in the same stress-response gene, WHI2. The WHI2 mutation rescues the mitochondrial respiratory defect (petite formation) caused by FIS1 deficiency, but also causes a failure to suppress cell growth during amino-acid deprivation. Thus, loss of Fis1 drives the selection for specific compensatory mutations that confer defective growth control and cell death regulation, characteristic of human tumor cells. The important long-term survival function of Fis1 that is compensated by WHI2 mutation appears to be independent of fission factor Dnm1/Drp1 and its adaptor Mdv1, but may be mediated through a second adaptor Caf4, as WHI2 is also mutated in a CAF4 knockout.

The recently introduced fast protein liquid chromatography (FPLC) system of Pharmacia (Uppsala, Sweden) was employed to isolate rather large peptides derived from thermolysin by selective chemical fragmentation at methionine in positions 120 and 205 of the polypeptide chain of 316 amino acid residues. Thermolysin was cleaved under conditions of limited fragmentation in order to produce, besides fragments 1 -1 20, 12 1 -205 and 206 -3 16, the overlapping fragments 1-205 and 121 -3 16. These polypeptides were separated employing prepacked Mono Q or Mono S columns (quaternary ammonium and sulfonic acid support, respectively). The columns were equilibrated with acetate-7 M urea buffer, pH 5.0 or 6.0, and eluted with a gradient of sodium chloride or acetate. Separations were achieved in 10-20min and were carried out also at a semipreparative level (1-3 mg per run). All five protein fragments were isolated in homogeneous form, as judged by amino acid analysis and electrophoresis. Considering that protein fragmentation with cyanogen bromide is the most commonly used procedure to achieve selective chemical fragmentation of a polypeptide chain, these results indicate that FPLC with ionic exchangers can be usefully employed to isolate rather large protein fragments especially suitable for automatic sequence analysis with the sequenator. Abbreviations used: CNBr, cyanopen bromide; BDTA, ethylcnediaminotetraacetic acid; FPLC, fast protein liquid chromatography; Mono Q, anion exchange liquid chromatography (HPLC) techniques, column containing a hydrophilic polymer carrying quaternary amino groups; Mono S , cation exchange which are now finding extensive use in the column containing a hydrophilic polymer carrying sulfonic groups; SDS sodium dodecyl sulfate; TCA-isolation and characterization of peptides and thermolysin, trichloroacetic acid-inactivated thcrmo-proteins -6). Because HPLC offers advantages lysin.

Molecular dynamics a b s t r a c t This study reports the comparative molecular modeling, docking and dynamic simulations of human a9a10 nicotinic acetylcholine receptors complexed with acetylcholine, nicotine and a-conotoxin RgIA, using as templates the crystal structures of Aplysia californica and Lymnaea stagnalis acetylcholine binding proteins. The molecular dynamics simulations showed that Arg112 in the complementary a10(À) subunit, is a determinant for recognition in the site that binds small ligands. However, Glu195 in the principal a9(+), and Asp114 in the complementary a10(À) subunit, might confer the potency and selectivity to a-conotoxin RgIA when interacting with Arg7 and Arg9 of this ligand.

The importance of comets for the origin of life on Earth has been advocated for many decades. Amino acids are key ingredients in chemistry, leading to life as we know it. Many primitive meteorites contain amino acids, and it is generally believed that these are formed by aqueous alterations. In the collector aerogel and foil samples of the Stardust mission after the flyby at comet Wild 2, the simplest form of amino acids, glycine, has been found together with precursor molecules methylamine and ethylamine. Because of contamination issues of the samples, a cometary origin was deduced from the (13)C isotopic signature. We report the presence of volatile glycine accompanied by methylamine and ethylamine in the coma of 67P/Churyumov-Gerasimenko measured by the ROSINA (Rosetta Orbiter Spectrometer for Ion and Neutral Analysis) mass spectrometer, confirming the Stardust results. Together with the detection of phosphorus and a multitude of organic molecules, this result demonstrates that c...

Bacterial surface proteins are important molecules in the infectivity and survival of pathogens. Surface proteins on gram-positive bacteria have been shown to attach via a transpeptidase, termed sortase, that cleaves an LPXTG sequence found close to the C termini of nearly all surface proteins on these bacteria. We previously identified a unique enzyme (LPXTGase) from Streptococcus pyogenes that also cleaves the LPXTG motif with a catalytic activity higher than that of sortase, suggesting that it plays an important role in the attachment process. We have now purified and characterized an LPXTGase from Staphylococcus aureus and found that it has both similar and unique features compared to the S. pyogenes enzyme. The S. aureus enzyme is glycosylated and contains unusual amino acids, like its streptococcal counterpart. Like the streptococcal enzyme, staphylococcal LPXTGase has an overrepresentation of amino acids found in the peptidoglycan, i.e., glutamine/glutamic acid, glycine, alan...

The aggregation of proteins or peptides in amyloid fibrils is associated with a number of clinical disorders, including Alzheimer's, Huntington's and prion diseases, medullary thyroid cancer, renal and cardiac amyloidosis. Despite extensive studies, the molecular mechanisms underlying the initiation of fibril formation remain largely unknown. Several lines of evidence revealed that short amino-acid segments (hot spots), located in amyloid precursor proteins act as seeds for fibril elongation. Therefore, hot spots are potential targets for diagnostic/therapeutic applications, and a current challenge in bioinformatics is the development of methods to accurately predict hot spots from protein sequences. In this paper, we combined existing methods into a meta-predictor for hot spots prediction, called MetAmyl for METapredictor for AMYLoid proteins. MetAmyl is based on a logistic regression model that aims at weighting predictions from a set of popular algorithms, statistically selected as being the most informative and complementary predictors. We evaluated the performances of MetAmyl through a large scale comparative study based on three independent datasets and thus demonstrated its ability to differentiate between amyloidogenic and non-amyloidogenic polypeptides. Compared to 9 other methods, MetAmyl provides significant improvement in prediction on studied datasets. We further show that MetAmyl is efficient to highlight the effect of point mutations involved in human amyloidosis, so we suggest this program should be a useful complementary tool for the diagnosis of these diseases.

Methunobacterium bryantii contains a single electrophoretically discernible superoxide dismutase, which constitutes 0.4% of the extractable protein. This enzyme has been purified to electrophoretic and ultracentrifugal homogeneity. It appears to be a tetramer. The subunits were tenaciously, but noncovalently bonded and were of identical size. The molecular weight of the enzyme was found to be '91,000 + 2000. The specific activity of this enzyme was identical to that previously noted for the corresponding enzyme from Escherichia co& The enzyme contained 2.7 atoms of Fe, 1.7 atoms of Zn, and less than 0.2 atoms Mn per tetramer. Its amino acid composition placed this enzyme with the other Mn-and Fe-containing superoxide dismutases. The M. bryantii enzyme was also similar to previously described Fe-containing superoxide dismutases in its optical and electron paramagnetic resonance spectra and in its susceptibility to inactivation by HzOz. The M. bryantii enzyme was ininhibited by NT, but was less sensitive towards this inhibitor than other iron-containing superoxide dismutases.

The effects on performance, digestibility, N utilization and plasma amino acid concentrations of dietary chickpea (Cicer arietinum, var. Kabuli) seed meal, globulin proteins or buffer-insoluble residue (starch / non-starch polysaccharides (NSP)/ lignin) were studied in growing rats. Chickpea meal, defatted soybean meal, chickpea globulins and lactalbumin were each incorporated into diets as the sole source of dietary protein (100 g/kg).

Muscle wasting and changes in muscle protein metabolism in chronic obstructive pulmonary disease. R.T. Jagoe, M.P.K.J. Engelen. #ERS Journals Ltd 2003. ABSTRACT: Loss of skeletal muscle mass is now recognised as an important feature of chronic obstructive pulmonary disease (COPD) which contributes to symptoms and influences prognosis. The changes in skeletal muscle remain poorly understood, largely because only a few studies have been performed to define the adaptations in whole body and muscle protein metabolism in COPD.

The lipopeptide antibiotic surfactin is a potent extracellular biosurfactant produced by various Bacillus subtilis strains. Biosynthesis of surfactin was studied in a cell-free system prepared from B. subtilis ATCC 21332 and OKB 105, which is a transformant producing surfactin in high yield [Nakano, M. M., Marahiel, M.

Centaurea solstitialis (yellow star thistle) has been proven to cause equine nigropallidal encephalomalacia in horses. Over the last fifty years, nigropallidal encephalomalacia has been of interest to human medicine due to the possible connection with Parkinson's disease. Previous studies indicated the presence of neurotoxic nitrogenous compounds in polar extracts of the plant. In order to give a more detailed description of the nitrogen-containing fraction of C. solstitialis, various samples were collected at different development stages. Different aliquots of the same aqueous extract were directly derivatized with ophthaldialdehyde and dansyl chloride and analyzed separately by reversed-phase HPLC. A complete profile of the free nitrogenous fraction of C. solstitialis was given and results obtained with the two derivatization procedures were compared. No particularly high level of free aspartic and glutamic acids, two potent neuroexcitotoxic amino acids, were found in polar extracts of the plant. Tyramine resulted to be the most important biologically active amine present in C. solstitialis (with a mean concentration of 2.0 mg/100 g of dry weight). q

An experiment with male turkeys was conducted to test the hypothesis that turkey production performance responds positively to extra crystalline amino acid supplementation (lysine, methionine, and threonine) when subjected to a high ambient temperature regimen (HT) in the grower period. Two diets were formulated to provide lysine, methionine, and threonine concentrations that either 1) met the breeder recommendations or 2) contained 10% higher lysine and methionine concentrations from 22 to 134 d of age and 10% higher threonine concentration from 22 to 68 d of age. Both diets were fed at two temperatures (15 or 25 C) from 42 d of age onward.

Acute intravenous amino acid infusion or a highprotein diet increases renal plasma flow and the glomerular filtration rate in healthy subjects. Conversely, a low-protein diet reduces renal plasma flow and glomerular filtration rate. The aim of this study was to investigate the effect of intravenous amino acid infusion and dietary proteins on kidney function in cirrhosis. Protocol 1: renal plasma flow and glomerular filtration rate were measured before and during intravenous administration of a 10% amino acid solution (0.043 ml/kg/min) to eight compensated cirrhotic patients (group l), nine nonazotemic cirrhotic patients with ascites (group 2) and seven cirrhotic patients with ascites and functional renal failure (group 3). Amino acid administration induced a significant increase in renal plasma flow and glomerular filtration rate in all groups studied. Renal plasma flow and glomerular filtration rate increased by 16% and 14%, respectively, in group 1; 31% and 22% in group 2 and 25% and 21% in group 3. Protocol 2: Renal plasma flow and glomerular filtration rate were measured in nine cirrhotic patients with ascites after 11 days on a low-protein diet (0.5 gm/kg body weight/day) and also after the patients followed for 11 days a moderately high-protein diet (1.5 g m / kg body weight /day). The moderately high-protein diet was associated with a significant increase in renal plasma flow (12%) and glomerular filtration rate (13%) compared with values obtained while the patients followed the low-protein diet. Plasma glucagon levels increased markedly during the intravenous administration of amino acid and the intake of the moderately highprotein diet. However, no correlation existed between changes in renal plasma flow and glomerular filtration rate induced by intravenous amino acid or dietary proteins and changes in plasma glucagon levels. The renal effects of intravenous amino acid or dietary pro-

Bacillomycin Lc, a newantifungal antibiotic of the iturin class, was isolated from a strain of Bacillus subtilis as a set of five congeners. The structure as determined by chemical and spectrometric analyses has been shown to differ from that of bacillomycin L by sequence changes from aspartate-1 to asparagine-1 and from glutamine-5 to glutamate-5. The five congeners differ from each other only in the structure of the aliphatic side chain of the constituent /?-amino acid. The hydrophobicity of the /?-amino acid affects the antifungal activity of the congener, as activity increased in the order of increased congener retention on a reversed-phase HPLCcolumn.

We have studied the distribution of the protein synthesis inhibitory activity in the tissues of Saponaria officinalis L. (Caryophyllaceae). Seven major saporins, ribosome-inactivating proteins, were purified to apparent homogeneity from leaves, roots and seeds using a new procedure of RIPs isolation including ion-exchange and hydrophobic chromatography. They all catalysed the depurination of rat liver ribosomes, which generate the Endo's diagnostic rRNA fragment upon treatment with acid aniline, thus indicating that m 4324 from the 28S rRNA has been released (Endo et al. (1987) J. Biol. Chem. 262, 5908-5912). The molecular mass of saporins by SDS-PAGE ranged between 30.2 and 31.6 kDa and by gel-filtration between 27.5 and 30.1 kDa. Amino acid composition and amino-terminal amino acid sequence indicate that all saporins may be considered isoforms. Only two saporins present in roots were glycosylated (SO-R1 and SO-R3). All saporins are very active on cell-free translation systems derived from rabbit reticulocyte lysates, rat liver, Triticum aestivum L., Cucumis sativus L. and Vicia sativa L. However, they are poor inhibitors of an Escherichia coli translation system. They inhibit protein synthesis in HeLa, BeWo and NB 100 cells, HeLa cells being the most resistant. The enzymatic activity of at least one saporin isoform was dependent on magnesium concentration in the standard rat liver cell-free system.

It is well recognized that variation in the geographical distribution of prevalence of multiple sclerosis (MS) exists: increasing the latitude its prevalence increases as well, but the underlying causes of such dissimilarity still remained elusive as of today. Currently, the most accredited hypothesis is that the closer to the equator the more pronounced is the amount of sunlight which, in turn, increases the production of vitamin D. Cholecalciferol is indeed deficient in MS patients, but this factor does not explain by itself the etiopathogenesis of the disease. In the present study, to search for a pattern and provide a model of the disease's etiology consistent with this regional factor, as well with its changing ethnic, sex-ratio, lifestyle variations and the other unexplained aspects of MS, an extensive analysis of peer-reviewed literature and data was conducted. The arisen hypothesis was that, increasing the latitude, the factor that varies and can have the stronger effect on the human organism, is the continuous and ever-increasing diversity of the natural light-dark cycle. The consequent effort of the suprachiasmatic nucleus to entrain the organism's circadian rhythm affects the hypothalamic-pituitary-adrenal axis resulting in desynchronizing the central and peripheral circadian clocks and pathologizing the immunitary system. To verify such hypothesis, a theoretical framework of the etiopathogenesis, coherent with the gathered literature, was conceived and a demonstration to corroborate it was eventually devised and performed. The results underscored that people living in countries subjected to a further circadian disruptive factor, as daylight saving time, have a 6.35 times higher prevalence of MS than States placed on their same latitude that do not observe it, thus strongly supporting the hypothesis. As further reinforcement of the conclusions, it is worth mentioning that the levels of polyamines rise abruptly in autoimmune diseases. Moreover, among their numerous roles, these polycations participate to the regulation of the circadian clock so their sudden variation might disrupt it. Following these interesting findings, new perspectives in therapies are, therefore, proposed.

A novel toxic cyclopeptide from Amanita suballiacea (Murr.) mushrooms that possesses structural features similar to viroidin is described. This peptide, alloviroidin, is identical with viroidin in mass, affinity for actin, and all amino acids except for one. The single discernible difference between the two peptides exists in the configuration at carbon 4 of the 4,5-dihydroxyleucine residues, as shown by a combination of chemical modification and magnetic resonance experiments. The configuration of this residue in viroidin is similar to that of phalloidin and is 2S,4R, while that in alloviroidin is established to be 2S,4S. This peptide is thus unique in its hydroxylation pattern among both the virotoxins and phallotoxins and may be an intermediate for more highly hydroxylated virotoxins, such as viroisin.

Lyophilization is a cost-effective method for biological specimen preservation but detailed tissue-specific reference protocols are still lacking. Moreover, data are limited on the long-term stability of proteins and nucleic acids in lyophilized samples.Here, we offer lyophilization protocols for various rat and mouse tissues (kidney, heart, liver, lung, aorta, and skin) coupled with technical hints for optimal sample preparation. We demonstrate that lyophilized samples stored at 4 °C for 20 months can yield protein and RNA of similar quantity and quality to −80 °C storage, while phosphorylated proteins are preserved as well. Freeze-dried and subsequently pulverized samples can provide more consistent, more reliable data especially when investigating focal injuries, such as fibrosis. We developed a protocol for the concentration of biological solutions and achieved 20-times concentration in human peritoneal dialysis effluent solution which enables the previously unattainable detecti...

Separation of globin chains by electrophoresis provides a simple and rapid method for the determination of the Gy/Ay ratio in human fetal haemoglobin, and of biosynthetic rates of the globin chains. Whole haemolysates were analysed by electrophoresis on polyacrylamide gels in urea, acetic acid and Triton X-100.

Background: Pseudomonas fluorescens SBW25 has been extensively studied because of its plant growth promoting properties and potential as a biocontrol agent. The genome of SBW25 has been sequenced, and among sequenced strains of pseudomonads, SBW25 appears to be most closely related to P. fluorescens WH6. In the authors' laboratories, WH6 was previously shown to produce and secrete 4-formylaminooxyvinylglycine (FVG), a non-proteinogenic amino acid with selective herbicidal and antimicrobial activity. Although SBW25 does not have the genetic capacity to produce FVG, we were interested in determining whether this pseudomonad might produce some other type of non-proteinogenic amino acid.

We describe an aerosol-liquid cycle that launches the autocatalytic amplification of any initial imbalance of the order of 10 ÿ7 % (1 ppb) up to total chiral purity in a single step process. Crystal nucleation of NaClO 3 is initiated at the aerosol air-water interface where, due to the accumulation of ambient chiral impurities or added hydrophobic chiral aminoacids in tiny concentrations (ppb), the initial levorotatory (l) and dextrorotatory (d) excess will not be produced with equal probability. The enantiomeric yield is then enhanced up to homochirality by recycling the crystallites through a liquid phase. In the absence of added catalysts this process leads to preferential (d) homochiral crystallizations in a ratio of 4:1 which is due to ambient contamination. By adding only 2 ppb of (L) or (D) Phe, we induce a final preferential homochiral crystallization of (d) or (l) handedness, respectively, in a ratio of 2:1.

Ready-for-use' instruments from surgical instrument trays were examined after routine cleaning and sterilization in a blinded study. These reprocessed instruments originated from five National Health Service hospital trust sterile service departments in England and Wales. Determination of residual protein and peptide contamination was carried out by acid stripping of the instrument surfaces, hydrolysis of the constituent amino acids and quantitative total amino acid analysis. One hundred and twenty instruments were analysed, and the median levels of residual protein contamination per instrument for the individual trays were 267, 260, 163, 456 and 756 mg. Scanning electron microscopy and energy dispersive X-ray spectroscopic analyses of the instruments showed that tissue deposits were localized on surfaces, but there was no significant correlation between overall protein soiling and instrument complexity. The highest levels of residual contamination were found on instruments used for tonsillectomy and adenoid surgery.

Radiopharmaceuticals are able to give functional information about systems, organs or cells. This functional information can be about different cell mechanisms or molecular pathways. In terms of systems or organs this information can be assessed through biodistribution studies while that in terms of cells can be evaluated through more detailed research, like calculation of influx and efflux indexes or binding studies.

The aim of this investigation was to analyze two samples of pollen from jandaíra stingless bees (Melipona subnitida) in view of their mineral composition, free amino acids and sugars. Palynological analysis showed that the predominant pollen was monofloral from Senna sp. species (94.5%, pollen 2011) and the second pollen sample showed the presence of two primary species, Chamaecrista sp. (39.2% pollen 2009) and Mimosa tenuiflora (43.5%) (pollen 2009). The highest mineral content was potassium. The bee pollen contained 20.8% and 31.0% of mannitol in samples from 2011 and 2009, respectively. Proline and serine are the predominant amino acids. High content of essential amino acids, minerals and the sugar mannitol confirmed high nutritional value of pollen samples from jandaíra bees.

For reversed phase separation amino acids are usually derivatized. Several derivatization reactions are carried out at basic pH. In the present work, influence of three basic buffer solutions on liquid chromatography electrospray ionization mass-spectrometric (LC-ESI-MS) analysis of amino acid derivatives was studied. Borate buffer -the most common derivatization buffer -was found to influence ESI ionization up to 23 min retention time. For 9-fluorenylmethylmethoxycarbonyl chloride (Fmoc-Cl derivatization) carbonate buffer should be preferred as it provides higher responses. Hexafluoroisopropanol (HFIP) buffer improves chromatographic peak shapes and responses for diethyl ethoxymethylenemalonate (DEEMM) derivatives.

Proteins that are highly expressed and composed of amino acids that are costly to synthesize are likely to place a greater drain on an organism's energy resources than proteins that are composed of ingested amino acids or ones that are metabolically simple to produce. Silks are highly expressed proteins produced by all spiders and many insects. We compared the metabolic costs of silks spun by arthropods by calculating the amount of ATP required to produce their component amino acids. Although a definitive conclusion requires detailed information on the dietary pools of amino acids available to arthropods, on the basis of the central metabolic pathways, silks spun by herbivorous, Lepidoptera larvae require significantly less ATP to synthesize than the dragline silks spun by predatory spiders. While not enough data are available to draw a statistically based conclusion, comparison of homologous silks across ancestral and derived taxa of the Araneoidea seems to suggest an evolutionary trend towards reduced silk costs. However, comparison of the synthetic costs of dragline silks across all araneomorph spiders suggests a complicated evolutionary pattern that cannot be attributed to phylogenetic position alone. We propose that the diverse silk-producing systems of the araneoid spiders (including three types of protein glues and three types of silk fibroin), evolved through intra-organ competition and that taxon-specific differences in the composition of silks drawn from homologous glands may reflect limited or fluctuating amino acid availability. The different functional properties of spider silks may be a secondary result of selection acting on different polypeptide templates.

Derivatives of the tyrosine kinase inhibitor dasatinib were synthesized by esterification with 25 carboxylic acids, including amino acids and fatty acids, thereby extending the drug to interact with more diverse sites and to improve specificity. The dasatinib-l-arginine derivative (Das-R, 7) was found to be the most potent of the inhibitors tested, with IC50 values of 4.4, <0.25, and <0.45 nm against Csk, Src, and Abl kinases, respectively. The highest selectivity ratio obtained in our study, 91.4 Csk/Src, belonged to compound 18 (Das-C10 ) with an IC50 value of 3.2 μm for Csk compared with 35 nm for Src. Furthermore, many compounds displayed increased selectivity toward Src over Abl. Compounds 15 (Das-glutamic acid) and 13 (Das-cysteine) demonstrated the largest gains (10.2 and 10.3 Abl/Src IC50 ratios). Das-R (IC50 =2.06 μm) was significantly more potent than the parent dasatinib (IC50 =26.3 μm) against Panc-1 cells, whereas both compounds showed IC50 <51.2 pm against BV-...

We present a complete DNA sequence and metabolic analysis of the dominant oral bacterium Fusobacterium nucleatum. Although not considered a major dental pathogen on its own, this anaerobe facilitates the aggregation and establishment of several other species including the dental pathogens Porphyromonas gingivalis and Bacteroides forsythus. The F. nucleatum strain ATCC 25586 genome was assembled from shotgun sequences and analyzed using the ERGO bioinformatics suite (http://www.integratedgenomics.com). The genome contains 2.17 Mb encoding 2,067 open reading frames, organized on a single circular chromosome with 27% GC content. Despite its taxonomic position among the gram-negative bacteria, several features of its core metabolism are similar to that of gram-positive Clostridium spp., Enterococcus spp., and Lactococcus spp. The genome analysis has revealed several key aspects of the pathways of organic acid, amino acid, carbohydrate, and lipid metabolism. Nine very-high-molecular-weight outer membrane proteins are predicted from the sequence, none of which has been reported in the literature. More than 137 transporters for the uptake of a variety of substrates such as peptides, sugars, metal ions, and cofactors have been identified. Biosynthetic pathways exist for only three amino acids: glutamate, aspartate, and asparagine. The remaining amino acids are imported as such or as di-or oligopeptides that are subsequently degraded in the cytoplasm. A principal source of energy appears to be the fermentation of glutamate to butyrate. Additionally, desulfuration of cysteine and methionine yields ammonia, H 2 S, methyl mercaptan, and butyrate, which are capable of arresting fibroblast growth, thus preventing wound healing and aiding penetration of the gingival epithelium. The metabolic capabilities of F. nucleatum revealed by its genome are therefore consistent with its specialized niche in the mouth. Fusobacterium nucleatum is a gram-negative anaerobe, which belongs to the Bacteroidaceae family and is found naturally in the microflora of the mouth in healthy or diseased humans. F. nucleatum is a very long rod with tapering ends and is one of the dominant species of the 500 or more organisms that coexist in the oral cavity (21). Many of the oral flora are commensals but a few are opportunistic pathogens. F. nucleatum can be isolated not only from the mouth but also from infections such as skin ulcers, peritonsillar abscesses, septic arthritis, and endocarditis (5). Several species of Fusobacterium have been isolated and studied, including F. necrophorum (the causative agent of Lemierre's syndrome), F. ulcercans (skin ulcers), F. russi (animal bite infections), and F. varium (eye infections), with F. nucleatum and F. necrophorum considered to be the most pathogenic. In the initial stages of the periodontal disease process, saccharolytic, aerobic Streptococcus spp. and other bacteria adhere to and colonize the tooth enamel and root surface. This sets the stage for F. nucleatum to coaggregate with these early colonizers and to permit late colonizers, including dental pathogens, to eventually form a biofilm. These complex interactions result in the release of factors that lead to tooth decay. Physical interaction is very specific among various genera in this complex microbial community. Due to the unusual length, adhesive nature, and other cell surface properties of F. nucleatum, periodontal disease-causing bacteria such as Porphyromonas gingivalis, Bacteroides forsythus, Actinobacillus actinomycetemcomitans, Treponema denticola, and Streptococcus spp. aggregate and thrive; hence, F. nucleatum is referred to as a "bridge bacterium" (5). The interaction between F. nucleatum and P. gingivalis has been reported to be very specific (19), mediated by a lactose-binding adhesin (20). The same adhesin protein mediates the binding of F. nucleatum to a variety of eukaryotic cell types including HeLa cells, buccal epithelial cells, macrophages, polymorphonuclear leukocytes, and gingival and periodontal ligaments (46). Helicobacter pylori was also shown to adhere selectively to F. nucleatum (1). Research on Fusobacterium spp. has focused primarily on species identification, oral ecology, cell-cell communication, extracellular surfaces, amino acid degradation, carbohydrate metabolism, organic acid fermentation, and antibiotic resistance (5, 21). However, many metabolic pathways and their roles in survival in such specialized niches are not known. Genetic experiments have been hampered by the lack of molecular tools, although recently three plasmids, pFN1, pFN2, and pFN3, have been isolated and sequenced from the strain ATCC 10953. A transformation system has been developed for gene manipulation using the pFN1 ori sequence (14). Although many potential virulence candidates have been

Background: Infants and children with chronic diarrhea (CD) often require specialized foods or parenteral nutrition (PN) to achieve adequate nutrient intakes to support growth and development. We assessed the efficacy of an amino acid-based formula (AAF) in supporting growth and improving symptoms in infants and children with CD from multiple etiologies. Methods: Two studies were conducted: CD study in children (CD-C) and CD study in infants (CD-I). Each was a single group, baseline-controlled study in which each subject served as his/her own control. At enrollment, all subjects had CD lasting > 2 weeks and had ≥ 4 stools/day. Subjects were fed an AAF for 80 days starting at SD5, and were assessed at SD 28 and 84. Results: CD-C: 18 of 19 subjects completed the study. At enrollment, the mean age was 5.6 ± 0.7 years, the most common diagnosis was short bowel syndrome (SBS) (n = 13), and 5 subjects with SBS were on PN. Subjects achieved significant increases in weight-forage z-scores (p = 0.026). Over 50% of subjects achieved improvements in clinical outcomes targeted most frequently by their physicians. Of the five subjects on PN at enrollment, four had substantial weight gain and four had their PN requirements decreased. CD-I: 22 of 27 subjects completed the study. At enrollment, the mean age was 3.3 ± 0.3 months, the most common diagnosis was food allergy (n = 20), and no subjects were on PN. Subjects achieved significant increases in weight-forage z-scores (p = 0.0023), significant decreases in the number of stools/day (p = 0.0012), and improvements in stool consistency (p = 0.0024). Over 80% of subjects achieved improvements in the clinical outcomes targeted most frequently by their physicians. Conclusions: Infants and children with CD fed an AAF for three months displayed significant improvements in weight-forage z-scores and clinical symptoms. Children dependent on PN also grew well and four of five decreased their dependence on PN. Trial registration: Both trials were registered on ClinTrials.gov (CD-C, NCT01812629; CD-I, NCT01820494).

Tissue free amino acid pools account for only a small fraction of the total amino acid content in the body. Blood or plasma free amino acid pools represent only a very small proportion of tissue free amino pools. Plasma free amino acid profiles have been used as a tool to study nutritional and metabolic aspects of protein status. Static measurements of plasma amino acid levels have limited value as sensitive indicators of protein status because such measurements do not reflect the magnitude of amino acid fluxes in and out free amino acid pools. Use of plasma amino acid profiles, under well defined experimental conditions, in assessing limiting amino acids and essential amino acid requirements in ruminants is discussed.

The mechanisms that control polyamine (PA) metabolism in plant cell lines with different embryogenic potential are not well understood. This study involved the use of two Araucaria angustifolia cell lines, one of which was defined as being blocked, in that the cells were incapable of developing somatic embryos, and the other as being responsive, as it could generate somatic embryos. Cellular PA metabolism was modulated by using 5 mM arginine (Arg) or ornithine (Orn) at two time points during cell growth. Two days after subculturing with Arg, an increase in citrulline (Cit) content was observed, followed by a higher expression of genes related to PA catabolism in the responsive cell line; whereas, in the blocked cell line, we only observed an accumulation of PAs. After fourteen days, metabolism was directed towards putrescine accumulation in both cell lines. Exogenous Arg and Orn not only caused a change in cellular contents of PAs, but also altered the abundance of a broader spectru...

Stability constants of binary and ternary complexes of Cu(II) and Zn(II) transition
metal ions withethambutol hydrochloride antitubercular drug (L) and serine(R1) valine(R2)
methionine (R3)glutamic acid(R4)amino acids(R) have been determined pH metrically at 300C
temperature and 0.1 M ionic strength(NaClO4) in aqueous solution. The formation of various
possible species has been evaluated by computer program and their stability were discussed in
terms of ∆LogK, Kr, KL and KR relative stability parameters.

An improved and efficient method for the determination of underivatized amino acids based on the use of CE coupled to evaporative light scattering detector (ELSD), involving carbon nanotubes, was successfully developed. Carboxyled single-walled carbon nanotubes were used for the first time to perform the clean-up of the analyzed samples, which were afterwards analyzed by CE-ELSD. White tea samples were used to demonstrate the usefulness of the CE-ELSD coupled methodology. A suitable interface, based on a triple tube design sprayer, was developed and successfully used for coupling both instruments. Parameters affecting the separation and determination, including the elimination of interferences, were studied and properly optimized. Under the optimized conditions good resolution was achieved for the separation of seven amino acids. The precision of the method, expressed as RSD, was found within the 3.5-5.3% range. The LOD obtained for the proposed method were in the 1.2-2.1 pg range and the LOQ, were in the 2.0-11.5 pg range, with injection pressure of 5 KPa for 20 s (15.3 nL). This method is simple, rapid, and selective compared with other conventional techniques.