bluuebunny/crossref_metadata_2025 huggingface dataset:
Extending Employability or Solving Employers' Recruitment Problems? Demand-led Approaches as an Instrument of Labour Market Policy
Authors: Gore Tony
Published: March 2005

For years, labour market policy in the UK has been dominated by activation measures that seek to increase employment levels among the unemployed. This supply-side focus emphasises individual characteristics and responsibilities in isolation from wider labour market factors. However, there is widening acceptance that policies for employability can provide a link between the supply and demand sides of the labour market. This implies the involvement of employers in the design of skills training and work experience programmes, and for these to be related to employment sustainability and career progression. Several projects in the UK under the New Deal Innovation Fund are currently testing the merits of what is termed a 'demand-led approach'. This paper examines this in terms of increasing job procurement and retention, and critically assesses the way in which it has been applied in the UK to date. It investigates the role played by labour market intermediaries in engaging both job seekers and employers, and considers the difficulties of successfully implementing the approach in different circumstances. It concludes that in most cases demand-led schemes in the UK have been less about improving employability than meeting employers' short-term labour needs.
Molecular Basis for the RIN4 Negative Regulation of RPS2 Disease Resistance
Authors: Day Brad , Dahlbeck Douglas , Huang Jeffrey , Chisholm Stephen T. , Li Donghui , Staskawicz Brian J.
Published: March 2005

Recent studies have demonstrated that RPS2, a plasma membrane–localized nucleotide binding site/leucine-rich repeat protein from Arabidopsis thaliana, associates with RPM1 Interacting Protein 4 (RIN4) and that this association functions to modulate the RPS2-mediated defense pathway in response to the bacterial effector protein AvrRpt2. In addition to negatively regulating RPS2 activity, RIN4 is also a target of AvrRpt2, a Cys protease and cognate bacterial effector protein of RPS2. Nicotiana benthamiana has been employed as a heterologous expression system to characterize the RPS2–RIN4 association, defining the domains in RIN4 required for the negative regulation of RPS2 activity. Upon inoculation of N. benthamiana leaves with Agrobacterium tumefaciens expressing RPS2, a rapid hypersensitive response (HR) is detected with 22 h of infiltration. The HR can be blocked by infiltrating the leaf with A. tumefaciens expressing RPS2 in the presence of RIN4, recapitulating the ability of RIN4 to interfere with RPS2-mediated resistance in Arabidopsis. Moreover, in the presence of RIN4, the RPS2-mediated HR can be restored by the delivery of AvrRpt2 via A. tumefaciens. This assay has been developed as a phenotypic marker for (1) the HR-inducing phenotype associated with RPS2, (2) negative regulation of RPS2 by RIN4, and (3) the AvrRpt2-mediated disappearance of RIN4. Here, we present a series of deletion and site-directed mutation analyses to identify amino acids in RIN4 required for the RPS2–RIN4 association and to distinguish these from residues in RIN4 that serve as a target sequence for AvrRpt2. In addition to characterizing the RPS2–RIN4 association in N. benthamiana, we have moved forward to show that the biological relevance of these amino acid changes is applicable in Arabidopsis as well. To this end, we have identified specific amino acids within the C-terminal half of RIN4 that are required for RPS2 regulation and association.
Regulation of root vole population dynamics by food supply and predation: a two‐factor experiment
Authors: Nie Haiyan , Liu Jike
Published: March 2005

This paper reports the effects of food supply, predation and the interaction between them on the population dynamics of root voles, Microtus oeconomus, by adopting factorial experiments in field enclosures. This two‐factor experiment proved the general hypothesis that food supply and predation had independent and additive effects on population dynamics of root voles. The experimental results proved the following predictions: (1) predation reduced population density and recruitment significantly; (2) food supply increased population density; (3) predation and food supply influenced spacing behavior of root voles separately and additively: Exposure to predation reduced long movements of root voles between trapping sessions; additional food supply reduced aggression level and home range size of root voles. Less movement of individuals that exposed to predators possibly reduced their opportunity of obtaining food and lessened population survival rate, which led population density to decrease. Smaller home range and lower aggression level could make higher population density tolerable. The interactive effect of predation and food on home range size was highly significant (P=0.0082<0.01). The interactive effect of food and predation on dispersal rate was significant (P<0.01). From the experimental results, we conclude that the external factors (predation, food supply) were more effective than internal factors (spacing behavior) in determining population density of root voles – under the most favorable external conditions (−P, +F treatment), the mean density and mean recruitment of root vole population was the highest; under the most unfavorable external conditions (+P, −F treatment), the mean density and mean recruitment of root vole population was the lowest.
Enhanced nitrogen loss may explain alternative stable states in dune slack succession
Authors: Adema Erwin B. , Van de Koppel Johan , Meijer Harro A. J. , Grootjans Ab P.
Published: March 2005

Ecological theory emphasizes competitive interactions between plant species when explaining primary succession in plants. Ecosystem processes, such as nutrient accumulation, are often regarded as independent, steering successional changes without being affected by the interacting plant species. We present experimental evidence that plant species in wet dune slack systems are able to affect ecosystem processes in their favor by reducing the rate of nitrogen accumulation by coupled nitrification–denitrification, promoting their competitive position. We compared denitrification rates of two early successional species having radial oxygen loss (ROL) with two non‐ROL late successional species in a mesocosm experiment. The denitrification rates were significantly higher in mesocosms planted with early successional species Littorella uniflora (P<0.001) and Schoenus nigricans (P<0.05), relative to the rates found in the presence of non‐ROL species, Carex nigra and Calamagrostis epigejos. We analyzed the consequences of enhanced nitrogen loss on the competition between an early and a late successional species by means of a simple theoretical model. Our analysis revealed that early successional species capable of ROL might retard successional changes and lock the ecosystem in an unproductive state for an extended period of time. We emphasize that understanding of ecosystem processes is crucial in successful conservation of high biodiversity vegetation in wet dune slacks.
Modelling epiphyte metapopulation dynamics in a dynamic forest landscape
Authors: Snäll T. , Pennanen J. , Kivistö L. , Hanski I.
Published: March 2005

We combine simulations with spatial statistics to estimate the parameters of a metapopulation model for the epiphytic lichenLobaria pulmonariaspecializing on aspen (Populus tremula) and goat willow (Salix caprea) in Fennoscandian boreal old‐growth forests. We estimated the parameters of a forest landscape model (FIN‐LANDIS) by repeatedly running simulations and selecting the set of parameters for tree ecology and fire regime that reproduced empirical host tree density and spatial patterns. Second, we tested which variables were important in epiphyte colonization and estimated the dispersal kernel. Third, we run a metapopulation model for the lichen across the estimated landscape scenarios and selected values for the remaining parameters that reproduced the empirical patterns of epiphyte occurrence. There was little variation in predicted dynamics, occupancy and spatial patterns between replicate metapopulation simulations. However, more data would be required for accurately estimating the parameters of FIN‐LANDIS primarily because of the inherent stochasticity in large scale forest fires. Following the beginning of fire suppression in the study area 150 years ago, the model predicts that lichen occupancy first increases but subsequently declines. The lower occupancy in the past than at present is explained by high rate of tree destruction by fires, which increases local extinction rate in patch‐tracking metapopulations. In the absence of fires, the occupancy increases because of lower extinction rate, but without forest fires or alternative means of host tree regeneration, the lichen is predicted to go ultimately extinct because of severely reduced density of aspen and goat willow.
Habitat‐dependent foraging in a classic predator–prey system: a fable from snowshoe hares
Authors: Morris Douglas W.
Published: March 2005

Current research contrasting prey habitat use has documented, with virtual unanimity, habitat differences in predation risk. Relatively few studies have considered, either in theory or in practice, simultaneous patterns in prey density. Linear predator–prey models predict that prey habitat preferences should switch toward the safer habitat with increasing prey and predator densities. The density‐dependent preference can be revealed by regression of prey density in safe habitat versus that in the riskier one (the isodar). But at this scale, the predation risk can be revealed only with simultaneous estimates of the number of predators, or with their experimental removal. Theories of optimal foraging demonstrate that we can measure predation risk by giving‐up densities of resource in foraging patches. The foraging theory cannot yet predict the expected pattern as predator and prey populations covary. Both problems are solved by measuring isodars and giving‐up densities in the same predator–prey system. I applied the two approaches to the classic predator–prey dynamics of snowshoe hares in northwestern Ontario, Canada. Hares occupied regenerating cutovers and adjacent mature‐forest habitat equally, and in a manner consistent with density‐dependent habitat selection. Independent measures of predation risk based on experimental, as well as natural, giving‐up densities agreed generally with the equal preference between habitats revealed by the isodar. There was no apparent difference in predation risk between habitats despite obvious differences in physical structure. Complementary studies contrasting a pair of habitats with more extreme differences confirmed that hares do alter their giving‐up densities when one habitat is clearly superior to another. The results are thereby consistent with theories of adaptive behaviour. But the results also demonstrate, when evaluating differences in habitat, that it is crucial to let the organisms we study define their own habitat preference.
The effect of food on microparasite transmission in the waterfleaDaphnia magna
Authors: Fels Daniel
Published: March 2005

Studies on dynamics of trophic transmission usually refer to the predation of animals that contain parasites. Here, in contrast, I study the transmission of external stages of parasites namely of spores of the microsporidian gut parasitesGlugoides intestinalisand Micro1. These spores are filter fed by the waterfleaDaphniamagna and I was particularly interested in the effect of food on transmission success. It was hypothesized that food has a double impact on transmission success because of increased spore uptake on the one hand and increased spore defecation of infected waterfleas on the other. I tested for these assumed food effects in exposure experiments where transmission could occur naturally from infected to uninfected waterfleas in the presence or absence of food.

The major assumptions were confirmed. In one system transmission success was increased fourfold in the presence of food and feeding infected hosts had lower spore loads. The other system showed evidence for these effects. Feeding activity correlated positively with food in both systems and spore loads in infected waterfleas partially determined spore loads in exposed waterfleas.

I discuss the impact of food on infected and uninfected waterfleas in particular and on transmission success and epidemiology in general.
Phosphatidylcholine‐specific phospholipase C but not gamma interferon regulate gene expression and secretion of CC Chemokine Ligand‐2 (CCL‐2) by human astrocytes during infection by Toxoplasma gondii
Authors: Durand François , Brenier‐Pinchart Marie‐Pierre , Berger Francois , Marche Patrice N. , Grillot Renée , Pelloux Hervé
Published: March 2005

We have used human astrocytoma‐derived cells to investigate the cellular responses of central nervous system cells to Toxoplasma gondii infection. At 24 h post inoculation, the secretion of CCL‐2 (or Monocyte Chemotactic Protein‐1) was augmented six‐fold over the control. This secretion was down‐regulated by D609, a specific inhibitor of phosphatidylcholine‐dependent phospholipase C (PC‐PLC), but not modulated by gamma interferon (IFN‐γ). Ribonuclease protection assay analyses showed significant down‐regulation of CCL‐2 mRNA production during infection by Toxoplasma gondii when cells were treated by D609. The mRNA levels of the seven other chemokines studied were not modified by D609. CCL‐2 seems to contribute to the cell recruitment during human cerebral reactivation of Toxoplasma gondii. Cellular production of this CC chemokine during toxoplasmosis may be regulated by a PC‐PLC‐dependent pathway.
Inflammatory antigens of Brugia malayi and their effect on rodent host Mastomys coucha
Authors: Dixit S. , Gaur R. L. , Khan M. A. , Saxena J. K. , Murthy P. S. R. , Murthy P. K.
Published: March 2005

The study was aimed at identifying pro‐ and anti‐inflammatory cytokine releasing potential of Brugia malayi adult worm fractions and their role in filarial infection and pathogenesis. THP‐1 cells were incubated with soluble somatic Brugia malayi adult worm extract (BmAS) and its Sephadex G‐200 fractions BmAFI, BmAFII and BmAFIII and the effect of the fractions on parasitological, immunological and lymph node parameters was assessed in Mastomys coucha. BmAFII stimulated the pro‐inflammatory TNF‐α, IL‐1β and IL‐6 release; IL‐10 release was insignificant. Sensitization of animals with BmAFII and subsequent intraperitoneal implantation of worms enhanced CMI response. BmAFII also increased lymph node weight and cellularity, stimulated lymph node mast cells and eliminated intraperitoneally instilled worms. BmAFI stimulated several folds more release of IL‐10, whereas TNF‐α release was negligible. Sensitization with BmAFI elicited low CMI responses, moderately stimulated mast cells and facilitated survival of implanted adult parasites. Fifty percent of naive animals exposed to BmAFI showed oedematous lymph nodes and increased node weight. NCP‐bound molecules corresponding to BmAFI and II showed cytokine‐stimulating potential in vitro. It is concluded that BmAFII is protective and stimulates pro‐inflammatory cytokines, whereas BmAFI facilitates parasite survival and stimulates IL‐10.
Recombinant SSP4 protein from Trypanosoma cruzi amastigotes regulates nitric oxide production by macrophages
Authors: Ramos‐Ligonio A. , López‐Monteon A. , Talamás‐Rohana P. , Rosales‐Encina J. L.
Published: March 2005

Acute infection with Trypanosoma cruzi is characterized by immunosuppression mediated by T cells and macrophages (Mφs). Nitric oxide (NO) production during the initial phase of acute infection might participate in the clearance of parasites by Mφs, whereas its overproduction during the late phase of acute infection would account for the immunosuppression observed. Trypanosoma cruzi molecules that might regulate the host responses have not been fully identified. Here, we demonstrate that active immunization with MBP::SSP4, a recombinant protein derived from a surface antigen specific of T. cruzi amastigotes (TcSSP4), was able to stimulate Ab production (IgG1, IgG2a, and IgG2b). On the other hand, MBP::SSP4 was able to stimulate NO production by peritoneal Mφs from BALB/c mice and Mφs from the J774 cell line. This effect was also observed at the level of inducible nitric oxide synthase (iNOS) detected by Western Blot. Furthermore, MBP::SSP4 was also shown to induce the expression of IL‐1α, IL‐6, IL‐12, IFN‐γ, and TNF‐α in normal animals, and IL‐10 in immunized animals. In addition the protein MBP::SSP4 was able to bind to the surface of PMφs and J774 Mφs. These results suggest that TcSSP4 could modulate Mφ NO production and this may represent a mechanism participating in the immunoregulatory processes during Chagas’ disease.
Potential transmission of West Nile virus in the British Isles: an ecological review of candidate mosquito bridge vectors
Authors: Medlock J. M. , Snow K. R. , Leach S.
Published: March 2005

Abstract. West Nile virus (WNV) transmitted by mosquitoes (Diptera: Culicidae) infects various vertebrates, being pathogenic for birds, horses and humans. After its discovery in tropical Africa, sporadic outbreaks of WNV occurred during recent decades in Eurasia, but not the British Isles*. WNV reached New York in 1999 and spread to California by 2003, causing widespread outbreaks of West Nile encephalitis across North America, transmitted by many species of mosquitoes, mainlyCulexspp. The periodic reappearance of WNV in parts of continental Europe (from southern France to Romania) gives rise to concern over the possibility of WNV invading the British Isles.

The British Isles have about 30 endemic mosquito species, several with seasonal abundance and other eco‐behavioural characteristics predisposing them to serve as potential WNV bridge vectors from birds to humans. These include: the predominantly ornithophilicCulex pipiensL. and its anthropophilic biotypemolestusForskål; tree‐hole adaptedAnopheles plumbeusStephens; saltmarsh‐adaptedOchlerotatus caspiusPallas,Oc. detritusHaliday andOc. dorsalis(Meigen);Coquillettidia richiardiiFicalbi,Culiseta annulataSchrank andCs. morsitans(Theobald) from vegetated freshwater pools;Aedes cinereusMeigen,Oc. cantansMeigen andOc. punctorKirby from seasonal woodland pools. Those underlined have been found carrying WNV in other countries (12 species), including the rarer British speciesAedes vexans(Meigen),Culex europaeusRamos et al.,Cx. modestusFicalbi andOc. sticticus(Meigen) as well as theAnopheles maculipennisMeigen complex (mainlyAn. atroparvusvan Thiel andAn. messeaeFalleroni in Britain). Those implicated as key vectors of WNV in Europe are printed bold (four species).

So far there is no proof of any arbovirus transmission by mosquitoes in the British Isles, although antibodies to Sindbis, Tahyna, Usutu and West Nile viruses have been detected in British birds. Neighbouring European countries have enzootic WNV and human infections transmitted by mosquito species that are present in the British Isles. However, except for localized urban infestations ofCx. pipiensbiotypemolestusthat can be readily eliminated, there appear to be few situations in the British Isles where humans and livestock are exposed to sustained risks of exposure to potential WNV vectors. Monitoring of mosquitoes and arbovirus surveillance are required to guard the British Isles against WNV outbreaks and introduction of more anthropophilic mosquitoes such asStegomyia albopicta(Skuse) andOchlerotatus japonicus(Theobald) that have recently invaded Europe, since they transmit arboviruses elsewhere.
Fluorometric Detection of Histidine‐Tryptophan Complexes in Peptides and Proteins
Authors: Shinttzky M. , Goldman R.
Published: March 2005

A study of the fluorescence properties of cyclo(l‐histidine‐l‐tryptophan), cyclo(l‐histidine‐d‐tryptophan), α,N‐acetyl‐l‐histidine‐l‐tryptophan methyl ester, α,N‐acetyl‐L‐histidine‐L‐tryptophan and (l‐histidine‐l‐tryptophan)3 has revealed an intramolecular interaction between the indole and the protonated imidazole side chains which results in marked quenching of the fluores‐cence intensity of the indole moiety. All five model compounds were found to possess a characteristic fluorometric titration curve which practically coincides with the potentiometric titration curve of the imidazole side chain of the histidine residue. Theoretical derivations of the pK from both titration curves are given. The extent of quenching by the protonated imidazole moiety, however, is different for each compound and varies from 40 to 80%.

To detect the presence of intramolecular histidine‐tryptophan complexes in natural peptides and proteins, fluorometric titrations in aqueous solutions were carried out in the range of pH 4–9. Marked changes in fluorescence intensity with pH in the range of pH 5 to 8.5, suggesting the existence of well defined complexes, were observed with papain, activated papain, chymopapain, ficin and bromelain. Glucagon, pepsinogen, pepsin and goose egg white lysozyme show no change in fluorescence in this pH range. In systems where a histidine‐tryptophan interaction was observed, the pK value of the interacting histidine was evaluated from the fluorometric titration curve.
The Molecular Properties of the Methaemoglobin of Chironomus plumosus L. in Solution
Authors: Behlke J. , Scheler W.
Published: March 2005

By means of an analytical ultracentrifuge the molecular weight of purified methaemoglobin from Chironomus plumosus has been determined at different pH values. Under weak acidic conditions (pH 5.2 to 7.0) the sedimentation constant amounts to 2.73 ± 0.02 S and the diffusion constant varies from 7.5 to 8.5 F. The molecular weight was calculated as 32,000 ± 1,500.

Outside of this pH‐range the molecular weight decreases. In alkaline solution (pH 9.2–10.0) the dimer molecule splits off reversibly into monomer subunits, with an average molecular weight of 14,600 ± 300.

In the pH‐range 7.5 to 9.0 and below 5.0 there exists a mixture of dimer and monomer molecules. The monomer‐dimer‐equilibrium is pH‐dependent. pH1/2 of this equilibrium coincides with the pH1/2 of the spectrophotometrically estimated main ionization constant (pH1/2= 8.3). The deprotonation of a haemin‐bound water molecule induces a change of the quarternary structure of the Chironomus‐methaemoglobin‐molecule.

According to the frictional ratio and the Stoke‐radius the monomer and the dimer MetHb‐molecules have an asymmetrical shape. Assuming a rotation ellipsoid (prolate type) for the monomer form (pH > 9.2) the axes are about 24 and 12.6 Å. The relatively high frictional ratio of the dimer molecule may be interpreted by an association of the α‐ and β‐monomers in direction of their longer axes.
On the Non‐Essentiality of Two Specific Disulphide Bonds in Ribonuclease for its Biological Activity
Authors: Neumann H. , Steinberg I. Z. , Brown J. R. , Goldberger R. F. , Sela M.
Published: March 2005

The reaction of phosphorothioate with bovine pancreatic ribonuclease at pH 9.0 in the absence of urea led to a derivative, denoted 4PS‐ribonuclease, in which two of the four disulphide bonds in the molecule were opened. 4PS‐ribonuclease is a unique molecular species, and not a mixture of native and open‐chain ribonuclease; this is apparent from electrophoretic studies. By the use of the “diagonal map” technique it has been established unequivocally that the disulphide bonds opened were between half‐cystines 3–8 and 4–5, counting from the amino terminus along the ribonuclease chain.

4PS‐ribonuclease is fully active enzymically toward RNA, and more active than the native enzyme toward cytidine 2′,3′‐cyclic phosphate. It is indistinguishable from native ribonuclease in its immunological reaction with antiserum to ribonuclease, and in its spectrophotometric titration, which shows that three of the six tyrosine phenolic groups in 4PS‐ribonuclease titrate abnormally. Unlike native ribonuclease, 4PS‐ribonuclease is digested by trypsin, even though at a lower rate than open‐chain ribonuclease.
Ferrochelatase Activity in Normal and Irradiated Animal Tissues
Authors: Goudy B. , Dawes E. , Wilkinson A. E. , Wills E. D.
Published: March 2005

Ferrochelatase activity is present in rat and mouse liver, heart, kidney, spleen, bone marrow and intestinal mucosa. Enzyme activity is concentrated in the mitochondrial fraction and absent from the microsomal fraction. About 20% of the total activity of liver is in the 100,000 ×g supernatant.

Formation of lipid peroxides in mitochondria leads to some loss of enzyme activity but extensive lipid peroxidation can occur without substantial reduction in enzyme activity.

Ferrochelatase in tissue homogenates, mitochondria or in a purified form is unaffected by 50 kilorads in vitro but is 15%–30% inhibited after keeping irradiated samples for 20 h at 4°.

In liver, heart and muscle homogenates the activity of ferrochelatase, in mice irradiated with a whole body dose of 850 rads was not outside the range of control values; but in kidney, spleen, bone marrow and in liver and kidney mitochondria, there was an increase of enzyme activity on the 3rd day after irradiation, followed by a decrease and return to normal on the 9th–10th days.

It is considered unlikely that a decline in ferrochelatase activity can account for the accumulations of iron found in tissues of irradiated animals or that it is an important site of radiation damage in vivo.
l‐Glutamate‐Phenylpyruvate Aminotransferase
Authors: Scandurra R. , Cannella C. , Ferretti M. G.
Published: March 2005

l‐glutamate‐phenylpyruvate aminotransferase was extracted from beef kidney mitochondria. The purification procedure included lysis of mitochondria in n‐butanol, ammonium sulphate fractionation between 55% and 80% saturation, gel filtration on G‐100 Sephadex and fractionation on DEAE‐Sephadex. The optimum pH of the enzyme is around 8. The enzyme is homogeneous to starch gel electrophoresis at various pH but shows two components when submitted to ultracentrifugal analyses.

A possible dissociation of the enzyme into subunits dependent on concentration is discussed.
Ribosomal Precursor RNA in Saccharomyces carlsbergensis
Authors: Retèl J. , Planta R. J.
Published: March 2005

Several lines of evidence for the occurrence of ribosomal precursor RNA in yeast are presented. This ribosomal precursor RNA has been found in the nuclear fraction, it is rapidly labeled with radioactive nucleotides and sediments faster than the two ribosomal RNAs. In addition another rapidly labeled RNA of high molecular weight has been found, which is turning over rapidly and is presumably of messenger nature. It proved to be possible to differentiate between both kinds of rapidly labeled RNA by using the technique of “shift‐up” and “shift‐down” in conjunction with pulse‐labeling.
Regulatory Effects of Nicotinamide on Tryptophan Pyrrolase Synthesis in Rat Liver in vivo
Authors: Cho‐Chung Y. S. , Pitot H. C.
Published: March 2005

Earlier studies had shown that the enzyme tryptophan pyrrolase which cat lyzes the first irreversible step in the conversion of tryptophan to functional nicotinyl derivatives in rat liver, was strongly inhibited by NADPH which may be considered the ultimate distal product in the conversion of tryptophan to derivatives of nicotinamide. As an extension of this work the present study has demonstrated that the administration of nicotinamide in vivo to adrenalectomized animals resulted in a strong suppression of the tryptophan‐mediated elevation of tryptophan pyrrolase but had little or no effect on the hormonal induction of this enzyme by corticosteroids. These effects of nicotinamide were in sofar specific as analogs of this compound had little or no effect on the specific tryptophan‐medidated elevation of tryptophan pyrrolase. Furthermore, the effect of nicotinamide was shown to be specific for the enzyme, tryptophan pyrrolase, having no effect on the tryptophan‐mediated increase in serine dehydratase. By means of anti‐tryptophan pyrrolase antibodies prepared in rabbits, the pulse labeling of tryptophan pyrrolase in vivo by [14C]phenylalanine showed that, in animals pre‐fed a 0% protein diet, tryptophan administration resulted in a doubling of the [14C]phenylalanine incorporation into tryptophan pyrrolase. Nicotinamide administration completely abolished this effect of tryptophan but had little effect on the dramatic stimulation of tryptophan pyrrolase synthesis by hydrocortisone. It is postulated that nicotinamide may exert its inhibitory effect on tryptophan induced synthesis of tryptophan pyrrolase at the level of genetic translation.
The Stereochemistry of α‐Oxidation of Fatty Acids in Plants
Authors: Hitchcock C. , Morris L. J. , James A. T.
Published: March 2005

Solutions of pea‐leaf acetone‐dried powder oxidise [1‐14C]palmitic acid to 14CO2, and oxidise [14C16]palmitic acid to 14CO2]pentadecanoic acid and [14C]myristic acid. If d‐2‐hydroxy[12C]‐palmitic acid is also present, these metabolites are slightly less radioactive; if l‐2‐hydroxy[12C]‐palmitic acid is also present, the metabolites are much less radioactive. It is concluded that the sequential α‐oxidation of long‐chain fatty acids involves the l‐2‐hydroxyacid preferentially, thus: RCH2CO2H→l‐RCHOHCO2H→RCO2H→etc.
Amino Acid Sequence around Disulfide Bridge of Pig Immunoglobin λ‐Chains
Authors: Franěk F. , Keil B. , Nvotný J. , ŠOrm F.
Published: March 2005

The λ‐chains of normal pig γG‐immunoglobulin were digested with pepsin in 1 M formic acid. Cystine‐containing peptides were isolated by gel filtration on Sephadex G‐25 and by chromatography on Dowex 50 and Dowex 1. The following structures were found:
image

In addition to these peptides obtained in a high yield similar peptides were found but their complete structures were not determined in view of their low yield. The structures found in the λ‐chains display a high degree of similarity with corresponding structures known to exist in human Bence‐Jones proteins of the l‐type.
Suppression of Pyruvate Oxidation in Liver Mitochondria in the Presence of Long‐Chain Fatty Acid
Authors: v. Jagow G. , Westermann B. , Wieland O.
Published: March 2005

An incubation system for rat liver mitochondria is described in which pyruvate is almost exclusively metabolised by oxidative decarboxylation. The oxidation of [1‐14C]pyruvate was determined by measurement of 14CO2‐formation and pyruvate disappearance. At the same time O2 consumption and ketone body formation were followed. Carnitine stimulated pyruvate oxidation. Oxidation of pyruvate was inhibited by physiological concentrations of palmitate. The inhibition was not relieved by increasing concentrations of DL‐carnitine. Palmitoyl‐carnitine inhibited pyruvate oxidation in the same manner as palmitate. The inhibition was not influenced by increasing the concentration of carnitine in the medium. The possible mechanisms for this effect of palmitate and palmitoyl‐carnitine on pyruvate oxidation are discussed. Under conditions of lack of carbohydrate the inhibition of mitochondrial pyruvate oxidation by fatty acid seems of major importance for directing pyruvate carbon into the direction of glucose synthesis.
A Collagenolytic Enzyme from Aspergillus oryzae
Authors: Nordwig A. , Jahn W. F.
Published: March 2005

A highly purified protease has been prepared from culture filtrates of Aspergillus oryzae. The process consists of four steps, namely the dialysis of the starting material, precipitation with acetone, separation by carrier‐free electrophoresis and fractionation with acetone. The specific activity of the protease is increased 40 times or more. The product was shown to be a single substance by means of the ultracentrifuge, disc electrophoresis and immuno‐electrophoresis. As far as investigated, it is free of non‐specific activities.

The molecular weight was found to be about 20,000 by elution from a Sephadex G‐75 column, by ultracentrifugation and by amino acid analysis. The pH optimum normally lies between 9 and 10. The protease cannot be activated with metal ions; it is, however, inhibited by Cu2+, Hg2+ and Zn2+ ions. It is also inhibited by di‐isopropyl phosphofluoridate and other compounds that react specifically with serine.

The protease splits denatured proteins preferentially, but not solely, at the peptide bonds of amino acids with acid side chains as shown by investigations on long‐chain model peptides. It also digests native collagen. The mode of splitting is the same as that by which the collagenase from Clostridium histolyticum splits the apolar regions. However, bonds involving hydroxyproline appear to be resistant to attack by the Aspergillus enzyme. In addition, the protease described here is capable of digesting bonds of the polar regions of the native collagen molecule. The protease also acts as an esterase.

On the basis of the various types of proteolytic specificity found for the protease a new concept is put forward for understanding collagen breakdown in vivo and in vitro.
Differences between the Reactivity of Endogenous and Exogenous Adenine Nucleotides in Mitochondria as Studied at Low Temperature
Authors: HELDT H. W. , KLINGENBERG M.
Published: March 2005

The rates of phosphorylation of endogenous and exogenous ADP and of the uncoupler stimulated hydrolysis of endogenous and exogenous ATP by rat liver mitochondria have been measured at temperatures between 2–14°.

1) The rate of the phosphorylation of endogenous ADP was found to be more than three times higher than the rate of the phosphorylation of exogenous ADP at 6°. The rates tend to approach each other at higher temperatures.

2) The rate of the carbonylcyanide phenylhydrazone stimulated dephosphorylation of endogenous ATP was more than twenty times higher than that of exogenous ATP below 6°. Even at room temperature there remains a considerable difference between the two rates. Hence, it is concluded that only the hydrolysis of endogenous ATP can be taken as a measure of the activity of mitochondrial ATPase. At low temperatures ATPase was not stimulated by dinitrophenol, and dinitrophenol caused an inhibition of carbonylcyanide phenylhydrazone stimulated ATPase.

3) Atractyloside inhibited only the reactivity of exogenous ADP and ATP, but had no influence on the rate of phosphorylation of endogenous ADP or the rate of carbonylcyanide phenylhydrazone stimulated hydrolysis of endogenous ATP.

4) In principle the same results were obtained with rat heart mitochondria, where the difference in the rate of reactivity of endogenous and exogenous adenine nucleotides was considerably smaller than in rat liver mitochondria.

It is concluded that the access of exogenous adenine nucleotides to the inner compartment of the mitochondria, where the endogenous adenine nucleotides are located and where oxidative phosphorylation takes place, is limited at low temperature by the adenine nucleotide translocation.
Bromthymol Blue as a pH Indicator in Mitochondrial Suspensions
Authors: MITCHELL P. , MOYLE J. , SMITH L.
Published: March 2005

The distribution of the functional group of the pH indicator bromthymol blue across the osmotic barrier component (or M phase) of the cristae membrane separating the inner from the outer aqueous phase of rat liver mitochondria has been measured by three partially independent methods. In mitochondria in State 5, under the usual conditions, only about one third of the total bromthymol blue reacts in the inner phase. In State 3, in presence of a high concentration of valinomycin in a KCI medium, approximately the same proportion of the bromthymol blue reacts in the inner phase as in State 5; but in State 4, practically all the bromthymol blue is expelled across the M phase and registers the pH of the outer phase.

The bromthymol blue technique is incompetent to measure the pH of the inner phase of mitochondria in State 4; but a method is described that will permit the approximate quantitative measurement of the intramitochondrial pH from the light extinction of bromthymol blue in State 5 (or in State 3 in presence of valinomycin) provided that the distribution of the indicator and the internal and external buffering powers of the mitochondrial suspension are known.

The expulsion of the bromthymol blue from the mitochondria in State 4 is attributable to the development of an electric potential across the M phase.
The Action of Phospholipase A on the Plasma Membranes of Rat Liver Cells
Authors: GALLAI‐HATCHARD J. , GRAY G. M.
Published: March 2005

Isolated liver cells were treated with snake venom phospholipase A to obtain information on the relative importance of different phospholipids in the plasma membrane to its structural stability.

The hydrolysis of phosphatidylethanolamine and lecithin by phospholipase A from Naia naia produced a change in the permeability of the liver cell plasma membrane which was arbitrarily assessed by measuring the amount of the intracellular enzyme glutamic oxaloacetic transaminase released into the incubation medium.

There was only a small leakage of glutamic oxaloacetic transaminase from the cell until 25–35% of the phosphatidylethanolamine had been hydrolysed. Further hydrolysis caused a rapid increase in the release of the enzyme. Very little lecithin was hydrolysed by the same enzyme concentrations.

After treatment with phospholipase A the cells were stained with either acridine orange, eosin or nigrosin and examined by light, phase contrast and fluorescent microscopy. Their gross morphology appeared to be unaltered by the hydrolysis of up to 50–60% of the cell phosphatidylethanolamine.

The results suggest that phosphatidylethanolamine is relatively more important than lecithin in maintaining, in terms of permeability to a protein molecule, the structural integrity of the liver cell plasma membrane.
Self‐Oscillations in Glycolysis 1. A Simple Kinetic Model
Authors: SEL'KOV E. E.
Published: March 2005

The paper describes a simple kinetic model of an open monosubstrate enzyme reaction with substrate inhibition and product activation. A comparison between the model and the phosphofructokinase reaction shows a close resemblance between their dynamical properties. This makes it possible to explain qualitatively most experimental data on single‐frequency oscillations in glycolysis. A mathematical analysis of the model has shown the following.

In the model, at a definite relationship between the parameters, self‐oscillations arise.

The condition of self‐excitation is satisfied more readily with a lower source rate, larger product sink rate constants, lower product‐enzyme affinity and higher enzyme activity.

Self‐oscillations exist only in a certain range of values of the parameter determining the degree of substrate inhibition. This range increases with decreasing source rate. Too strong or, conversely, too weak substrate inhibition leads to damped oscillations.

The period of self‐oscillations depends on the degree of substrate inhibition, the source rate, the sink rate constant, the enzyme activity, the affinity of the substrate and the product for the enzyme; it decreases with an increase in these values.

With an increase in the relative sink rate constant the steady state amplitude of self‐oscillations initially increases until a definite maximum is reached and then drops to zero.

A self‐oscillatory state in the phosphofructokinase reaction exists only when the maximum rate of this reaction is essentially higher than the source rate, and lower than the maximum rate of the reactions controlling the sink of the products.

An experimental investigation of self‐oscillations in the phosphofructokinase reaction may be considerably simplified by using a reconstituted system consisting of a small number of reactions with an irreversible sink of the products and artificial substrate supply. In this case the above relationship (section 6) should hold.
Purification and Properties of l‐6‐Hydroxynicotine Oxidase
Authors: DANG DAI Vo , DECKER K. , SUND H.
Published: March 2005

l‐6‐Hydroxynicotine oxidase, an inducible enzyme of nicotine catabolism in Arthrobacter oxidans, was obtained as a homogeneous protein, as judged by electrophoresis and ultracentrifuge studies; it forms hexagonal crystals in ammonium sulfate solution. Its molecular weight was determined to be 93,000. It contains two moles of FAD per mole of enzyme. In the presence of guanidine hydrochloride, enzymatically inactive components of molecular weight 47,000 were formed which appear to represent the subunits of the protein. No evidence for a participation of metal ions in the electron transport of the enzyme was obtained. Although Hg2+ and p‐chloromercuriphenylsulfonate strongly affect activity, N‐ethylmaleimide and 5,5′‐dithiobis‐(2‐nitrobenzoate) do not inhibit the enzyme. The latter compound reacts with one thiol group only after exposure of the enzyme to 7.5 M urea.
Nitration of Tyrosine Residues in the Pancreatic Trypsin Inhibitor with Tetranitromethane
Authors: MELOUN B. , FRIČ I. , ŠORM F.
Published: March 2005

Nitration of pancreatic trypsin inhibitor with tetranitromethane affords a mono‐and a disubstituted derivative which was shown by sequential analysis to contain nitrotyrosine in position 10 and in positions 10 and 21, respectively. Both retain full trypsin inhibitor activity. Measurements of optical rotatory dispersion in the 230 nm region indicate that the secondary structure of the protein is unaffected by nitration. The nitrated derivatives exhibit a conformation‐dependent side‐chain Cotton effect in the region of nitroaromatic absorption. No such Cotton effect is shown by a nitrotyrosine‐containing pentadecapeptide isolated from the tryptic digest from the nitrated and oxidized inhibitor.
Nouvelle méthode de purification des polypeptide synthétases VII
Authors: FISCHER‐FERRARO C. , BELJANSKI M.
Published: March 2005

A new method for purification of “polypeptide synthetase” from Alcaligenes faecalis is described in which nucleic acids are removed by purification on DEAE‐cellulose with a recovery of 60–75% of the initial enzymatic activity.

The enzyme preparation catalyses the binding of the naturally occuring l‐amino acids to a messenger‐like RNA whose properties have been previously described. The reaction requires the addition of any one of the four nucleoside triphosphates (ATP, GTP, CTP or UTP), which is degraded to the corresponding nucleoside diphosphate and orthophosphate. For certain amino acids, however, the requirement for a nucleoside triphosphate is limited to two of the four.

The purified enzyme preparation, unlike crude extracts, does not catalyse an amino dependent ATP‐[32P]PPi exchange. Neither crude extracts, nor purified preparations show detectable exchange between [32P]PPi and GTP, CTP, or UTP in the presence or in the absence of l‐amino acids.

The presence of more than one enzyme in the purified preparation was demonstrated by further substantial fractionation of [14C]leucine binding activity and separation of this from [14C]alanine binding activity. However, no fractionation, which was dependent on a specific nucleoside triphosphate, could be achieved.
Biochemical Studies on Lipopolysaccharides of Salmonella R Mutants
Authors: DRÖGE W. , LÜDERITZ O. , WESTPHAL O.
Published: March 2005

From Salmonella R lipopolysaccharides of the chemotypes Rc, Rd1, and Rd2 oligosaccharides were prepared and their structure studied by the following methods: quantitative sugar analyses; periodate oxidation followed by borohydride reduction in order to transform the l‐glycero‐d‐mannose residues into d‐mannose residues; methylation of original and transformed oligosaccharides, methanolysis and gas chromatographic identification of the methylated heptoses or mannoses; the action of glycosidases; serological tests. The results of the study are in agreement with the formulae given below for the different oligosaccharides. The anomeric form of the glucose unit linked to heptose, as well as that part of the oligosaccharides which contains the 2‐keto‐3‐deoxy‐octonate residues [KDO] remain uncertain.

Chemotypes Rd2: Heptose pα 1→[KDO]x; Rd1: Heptose pα 1→3 Heptose pα 1→[KDO]x; Re: Glucose 1→3 Heptose pα 1→3 Heptose pα 1→[KDO]x.
Biochemical Studies on Lipopolysaccharides of Salmonella R Mutants
Authors: MÜHLRADT P. , RISSE H. J. , LÜDERITZ O. , WESTPHAL O.
Published: March 2005

In an in vitro system, containing 20,000 ×g cell wall particles and 100,000 ×g cell sap supernatant of various Salmonella rough mutants the rate of galactose transfer from UDP‐[14C]‐galactose to the core lipopolysaccharide was studied and the following results obtained:

Cell wall partieles from the S. minnesota mutants mR5, mR7, and mR8 are poor acceptors for galactose. The lipopolysaccharides of these mutants contain heptose and phosphate in a molar ratio of 2:2, whereas in normal lipopolysaccharide of Salmonella the ratio is 2:3.

20,000 ×g cell wall fractions from mutants which produce lipopolysaccharide of normal phosphate content, such as those from S. minnesota mRz or S. typhimurium tmM are good acceptors for galactose.

Phosphate‐poor cell wall particles can be treated with ATP and enzyme extracts of mutants containing lipopolysaccharide of normal phosphate content to yield proper acceptors for galactose.

A radioactive disaccharide which chromatographically behaves like melibiose, and which after treatment with α‐galactosidase yields [14C]galactose can be isolated from partial hydrolysates of cell wall particles which have previously been incubated with UDP‐[14C]galactose and enzyme extract from tmM. This result is the same whether phosphate is already present in the cell wall fraction or is introduced enzymatically by the addition of ATP to the incubation mixture.

It is concluded that mR5 and mR8 are defective in a lipopolysaccharide phosphorylating enzyme and that lack of phosphate groups in the lipopolysaccharide results in an incomplete core because unphosphorylated lipopolysaccharide acts only as a poor acceptor in the galactose transfer reaction.
The Stereochemistry of α‐Oxidation of Fatty Acids in Plants
Authors: MORRIS L. J. , HITCHCOCK C.
Published: March 2005

When [1‐14C]palmitic acid is incubated with young pea leaves, or solutions of their acetonedried powders, d‐2‐hydroxy[1‐14C]palmitic acid accumulates. When l‐[1‐14C, 2‐3H]palmitic acid is similarly oxidised, d‐2‐hydroxy[1‐14C]palmitate is formed which retains the tritium (99% retention). When d‐[1‐14C, 2‐3H]palmitic acid is similarly oxidised, d‐2‐hydroxy [1‐14C]palmitate is formed with loss of tritium (24% retention). The mechanism of α‐hydroxylation is therefore replacement of α‐hydrogen by hydroxyl with overall retention of configuration.
Enzymatic Inactivation of Glutamine Synthetase in Enterobacteriaceae
Authors: GANCEDO C. , HOLZER H.
Published: March 2005

The inactivation of glutamine synthetase in vivo on addition of ammonium ions to the culture medium was studied in different microorganisms. Only those belonging to the family of Enterobacterioceae showed inactivation. The transfer activity of the enzyme was much less affected than the synthetic one. As in Escherichia coli, the glutamine synthetase of Salmonella typhimurium could be inactivated in vitro in a system requiring glutamine, ATP, Mg2+ and an enzyme.

The inactivating enzymes of E. coli and S. typhimurium have been separated and cross‐tested against their synthetases. The inactivating enzymes inactivated both glutamine synthetases in the in vitro inactivation process. In contrast, synthetases of microorganisms which did not exhibit in vivo inactivation were not affected by these inactivating enzymes.

The fact that only glutamine synthetases of Enterobacteriaceae could be inactivated suggests that this regulatory mechanism is limited to a group of organisms possibly derived from the same ancestor.