Stereospecific assignment of the NH2 resonances from the primary amides of asparagine and glutamine side chains in isotopically labeled proteins

An HMQC-based pulse scheme is presented for the stereospecific assignment of asparagineand glutamine side-chain amide protons. The approach makes use of the recently developedquantitative-J correlation spectroscopy [Bax, A. et al. (1994) Methods Enzymol., 239,79–105] to distinguish the E and Z primary amide protons and, as such, eliminates theneed for assignments derived from more time-consuming and potentially ambiguous NOEmethods. An application of this method to a uniformly 15N,13C-labeled cellulose-bindingdomain is presented. When used in combination with a NOESY-HSQC experiment, thepredominant 2 dihedral angles of two asparagine side chains in this protein can also bedefined.     

Mapping of a Gene for Long QT Syndrome to Chromosome 4q25-27

Long QT syndrome (LQTS) is a heterogeneous inherited disorder causing syncope and sudden death from ventricular arrhythmias. A first locus for this disorder was mapped to chromosome 11p15.5. However, locus heterogeneity has been demonstrated in several families, and two other loci have recently been located on chromosomes 7q35-36 and 3p21-24. We used linkage analysis to map the locus in a 65-member family in which LQTS was associated with more marked sinus bradycardia than usual, leading to sinus node dysfunction. Linkage to chromosome 11p15.5, 7q35-36, or 3p21-24 was excluded. Positive linkage was obtained for markers located on chromosome 4q25-27. A maximal LOD score of 7.05 was found for marker D4S402. The identification of a fourth locus for LQTS confirms its genetic heterogeneity. Locus 4q25-27 is associated with a peculiar phenotype within the LQTS entity.     

Localization of the CDKN4/p27Kip1 gene to human chromosome 12p12.3

CDKN4/p27Kip1 is a cyclin-dependent kinase (Cdk) inhibitor implicated in G1 phase arrest, which negatively regulates G1 phase progression in response to TGF, and might represent a tumor suppressor gene. We report here the chromosomal assignment of the human CDKN4 gene to chromosome 12p12.3 in close proximity to highly polymorphic microsatellite markers.     

Evolution of biological regulation networks under complex environmental constraints

This work investigates the influence of environmental inducers on the organization of cell regulation networks, using a connectionist approach. Protein interactions are modeled by an asymmetrical recurrent network, the units of which take continuous values. In contrast to classical models, we explicitly introduce a genome to encode the architecture of the system. This feature enables us to introduce an evolution model, in which a genetic algorithm that mimics the effects of evolution on proteins mutual interactions is used. We assume an efficient system to respond to persistent environmental stimuli, independently of their amplitude. Results are presented that show a structuration of the network with the emergence of specialized hierarchical structures. These structures seem to drive the system at the edge of chaos, so that it can present adapted responses to significant environmental changes.     

Inhibition of protein synthesis in a polyribosome-dependent cell-free system by a specific ribonuclease prepared from the follicles of two different stages of development of the silkmothAntherea pernyi

Summary Crude, cell-free protein-synthesizing systems were prepared from follicles of two different stages of development in the ovariole of the silkmothAntherea pernyi. The efficiency of the translation of natural and synthetic mRNAs in these systems was compared with that in a cell-free wheat germ system. A postmitochondrial extract (S-30) from the follicles almost completely inhibited protein synthesis in a polyribosome-dependent, cell-free systems. A specific ribonuclease, obtained from the post mitochondrial extract by ammonium sulphate precipitation, heat denaturation and DEAE-cellulose chromatography, inhibited polyribosome-dependent protein synthesis. The effect of this specific ribonuclease on the structural integrity of radioactive RNAs and ribosomal subunits, which were isolated from Ehrlich ascites tumor cells, was also studied.     

Characterization of temperate phages ofBacillus subtilis

Nine known temperature phages ofBacillus subtilis, including four that are newly isolated (ϱ6, ϱ10, ϱ14, and ϱ18), have been compared. Analysis by serology, immunity, host range, and adsorption site similarity place the phages into four groups: Group I, ϕ105, ϱ6, ϱ10, and ϱ14, which are 80–90% related; Group II, SPO2; Group III, ϕ3T and ϱ11, 100% related; and Group IV, SP16. The phage ϱ18 is largely uncharacterized, but is heteroimmune to other groups.     

Properties of polyphenol oxidase from tubers of the yam Dioscorea bulbifera

The subunit MW of Dioscorea bulbifera polyphenol oxidase (MW 115 000 ± 2000) determined by SDS-PAGE is ca. 31 000 indicating that the enzyme is an oligomeric protein with four subunits. K_i values of various inhibitors and their modes of inhibition have been determined with catechol and pyrogallol as substrates. p-Nitrophenol, p-cresol, quinoline and resorcinol are competitive inhibitors of catechol binding while only orcinol and p-nitrophenol behave in the same way towards pyrogallol as substrate. From the effect of pH on V_max, groups with pK values ca. 4.7 and 6.8 have been identified to be involved in catalytic activity. The Arrhenius activation energy (E_a) at pH 4.0 is 8.9 kcal/mol between 40–65°. At pH 7.0, the value is 22.1 kcal/mol between 40 and 60°. The enthalpies (ΔH) at pH 4.0 and pH 7.0 are 2.3 kcal/mol and 32.4 kcal/mol respectively. The results are discussed considering the conformational changes of the enzyme during substrate binding.     

Effects of Feeding and Drinking on Acetylcholine Release in the Nucleus Accumbens, Striatum, and Hippocampus of Freely Behaving Rats

Extracellular levels of acetylcholine (ACh) were measured in the nucleus accumbens (NAC), striatum (STR), and hippocampus (HIPP) using microdialysis in 30-min intervals before, during, and after free-feeding in 20-h food-deprived rats. The effects on ACh in the NAC and STR were also observed in response to water intake in 20-h water-deprived animals. Neostigmine was used in the perfusate to improve ACh recovery. Basal ACh was sensitive to tetrodotoxin and low calcium, and therefore largely neuronal in origin. Feeding caused a 38% increase in extracellular ACh in the NAC and no change in the STR or HIPP. Dopamine was also increased in the NAC (48%) and to a lesser extent in the STR (21%) following feeding. Drinking caused 18-20% increases in ACh release in both the NAC and STR. In a separate experiment, ACh release in the NAC was monitored in 10-min intervals during free-feeding; ACh increased in the interval immediately following maximal food intake. These results suggest a site-specific increase in ACh release following feeding that cannot be solely attributed to the activation associated with this behavior.     

Experimental infections with Nematodirus spathiger in rabbits.

The feasibility of infecting laboratory rabbits experimentally with the ovine nematode Nematodirus spathiger was examined. Eight-week-old rabbits were dosed either with 5000 or with 17,000 third-stage larvae and killed on days 10, 21 or 42 post-infection. With the lower dose, 20 to 40% of the inoculum were recovered at necropsy. Similar values were observed with the 17,000 dose on days 10 and 21 post-infection, but on day 42 the worm population was residual (0.6%). With both dose levels, during the course of infection, the worm populations were mainly composed of fourth-stage larvae and worm egg excretion was low. N. spathiger mainly inhabited the proximal jejunum. The results were compared with N. spathiger infection in sheep to assess the usefulness of the rabbit as an experimental model for Nematodirus infection.     

Membrane-associated myo-inositol dehydrogenase activity ofBacillus pumilus strain 5

Summary The influence of the microenvironment on the metabolic behaviour of immobilized cells is discussed in relation to literature data. A model is proposed in which decreased water activity and/or oxygen deficiency result in changed yields and/or new metabolic behaviour of the immobilized cells.