A T cell nuclear factor resembling NF-AT binds to an NF-kappa B site and to the conserved lymphokine promoter sequence "cytokine-1".

Nuclear extracts from a nontransformed murine T lymphocyte clone contained two inducible factors that bound to a nuclear factor kappa B (NF-kappa B) site. One factor was NF-kappa B, and the other was differentiated from NF-kappa B by its mobility in the electrophoretic mobility shift assay and its lack of sensitivity to protein kinase C depletion. Competition and methylation interference assays showed that the binding site for the novel factor was limited to nucleotides in the 3' half of the kappa B site. This part of the kappa B site resembled sequences in the binding site for a second inducible nuclear factor of T cells, NF-AT, as well as a conserved sequence found in several lymphokine genes, termed "cytokine-1" (CK-1). Competition and methylation interference analysis showed that both NF-AT and CK-1 sequences bound a factor similar to the novel kappa B-binding factor and that binding involved a four-nucleotide sequence (TTCC) that the kappa B, CK-1, and NF-AT sites have in common. The complexes that form with each site have characteristics of NF-AT: they are induced upon T cell receptor stimulation, are sensitive to protein synthesis inhibitors and cyclosporin A, and are not sensitive to protein kinase C depletion. Thus, a factor or factors similar to NF-AT can bind to three distinct promoter sequences which occur commonly in several T cell activation genes. These results raise the possibility that related factors binding to kappa B, CK-1, and NF-AT sequences could play a role in the coordinate induction of T cell activation genes. In addition, our results suggest that kappa B and CK-1 sites represent potential cyclosporin-sensitive promoter elements by virtue of their ability to bind an NF-AT-like factor.     

Activity and stability of yeast alcohol dehydrogenase (YADH) entrapped in aerosol OT reverse micelles

The activity and stability of yeast alcohol dehydrogenase (YADH) entrapped in aerosol OT reverse micellar droplets have been investigated spectrophotometrically. Various physical parameters, e.g., water pool size, w(0), pH, and temperature, were optimized for YADH in water/AOT/isooctane reverse micelles. It was found that the enzyme exhibits maximum activity at w(0) = 28 and pH 8.1. It was more active in reverse micelles than in aqueous buffers at a particular temperature and was denatured at about 307deg;C in both the systems. At a particular temperature YADH entrapped in reverse micelles was less stable than when it was dissolved in aqueous buffer.     

Synthesis of oligosaccharides containing the X-antigenic trisaccharide (alpha-L-Fucp-(1----3)-[beta-D-Galp-(1----4)]-beta-D-GlcpNAc) at their nonreducing ends.

The "armed" methyl 2,3,4-tri-O-benzyl-1-thio-beta-L-fucopyranoside was reacted with "disarmed" phenyl O-(tetra-O-acetyl-beta-D-galactopyranosyl)-(1----4)-6-O-benzyl-2- deoxy-2-phthalimido-1-thio-beta-D-glucopyranoside in the presence of CuBr2-Bu4NBr complex to give phenyl O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1----4)-O- [(2,3,4-tri-O-benzyl-alpha-L-fucopyranosyl)-(1----3])-6-O-benzyl-2-deoxy -2- phthalimido-1-thio-beta-D-glucopyranoside (6) as a novel glycosyl donor. The glycosylating capability of 6 was further examined using N-iodosuccinimide-triflic acid as a reagent. This led to the synthesis of a tetrasaccharide and a pentasaccharide incorporating the X-antigenic structure represented by 6.     

Oxidative fermentation of calcium-magnesium lactate to calcium-magnesium acetate deicing salt

Summary The oxidation of Ca–Mg lactate to Ca–Mg acetate (CMA) deicing salt was studied in pure cultures ofAcetobacter pasteurianus, Gluconobacter cerinus orG. oxydans. Gluconobacter sp., which maintained a practically self-controlled pH reaction and did not overoxidize acetate, appear to be potentially important for CMA production.     

Shoot organogenesis and plant regeneration in mulberry (Morus bombycis Koidz): Factors influencing morphogenetic potential in callus cultures

Regeneration of multiple shoots via callus induction and organogenesis was achieved in mulberry (Morus bombycis). Pre-soaked internodal explants in 4.4–8.9 M benzyladenine (BA) formed callus on Linsmaier and Skoog's medium containing 2,4-dichlorophenoxyacetic acid (9.05 M), -naphthaleneacetic acid (2.85 M) and BA (2.2 M). Explants soaked for 48 to 72 h in low levels of BA produced loose and nodular callus that showed regeneration ability. Calluses developed adventitious shoot buds within 3–4 weeks on medium containing BA (8.9 M). Fifteen-week-old calluses developed fewer shoot buds than five-week-old calluses, indicating a decrease in morphogenetic potential with increasing duration of callus cultures. Semi-thin section microscopy was used to evaluate incapability of sustained regeneration. Development of normal shoot bud primordia, due to sub-surface reorganisation, was high in young calluses. The decline in the frequency of shoot bud primordia formation with callus ageing is due to reduced cell division activity in epidermal as well as sub-epidermal layers.     

A chemiluminescent assay for quantitation of beta-galactosidase in the femtogram range: application to quantitation of beta-galactosidase in lacZ-transfected cells.

An optimized chemiluminescent assay for beta-galactosidase using a chemiluminescent substrate AMPGD (3-(4-methoxyspiro[1,2-dioxetane-3,2'-tricyclo-[3.3.1. 1(3,7)]decan]-4- yl)phenyl-beta-D-galactopyranoside) is described. This assay is rapid and sensitive and can detect as little as 2 fg of beta-galactosidase. Its use for the quantitation of beta-galactosidase in cells transfected with lacZ-expressing vectors is described. It is possible to detect a single cell stably expressing lacZ by this technique.     

Bancroftian microfilariasis in association with pulmonary tuberculosis. Report of a case with diagnosis by fine needle aspiration

A 25-year-old man presented with clinical and radiologic features suggestive of pulmonary tuberculosis. Since the examination of Ziehl-Neelsen-stained sputum smears for acid-fast bacilli was repeatedly negative, a transthoracic fine needle aspiration biopsy was performed. Papanicolaou-stained smears of the aspirate showed microfilariae of Wuchereria bancrofti and a tuberculous exudate but no acid-fast bacilli or classic granulomas. Subsequent sputum samples did show acid-fast bacilli, while a nocturnal peripheral blood sample showed microfilariae.     

Purification and characterization of recombinant antistasin: a leech-derived inhibitor of coagulation factor Xa

Antistasin (ATS) is a selective, tight-binding inhibitor of blood coagulation Factor Xa originally isolated from the salivary glands of the Mexican leech Haementeria officinalis. In order to provide sufficient quantities of ATS to further investigate the role of Factor Xa in blood coagulation, a recombinant version of ATS has been produced in an insect baculovirus host-vector system. In this study, we describe the purification and in vitro and in vivo characterization of a single recombinant antistasin (rATS) isoform. The purified protein constitutes a minor isoform relative to the more abundant ATS isoforms present in leech salivary gland extracts. In vitro, rATS inhibits purified human Factor Xa stoichiometrically, prolongs plasma-based clotting assays at nanomolar concentrations, and like native ATS, is cleaved at a single position by Factor Xa during the course of inhibition. An initial evaluation of the in vivo efficacy of rATS was addressed utilizing a rhesus monkey model of mild disseminated intravascular coagulation. rATS was shown to fully suppress thromboplastin-induced fibrinopeptide A generation in a dose-dependent fashion. The availability of rATS should provide a valuable tool for the critical evaluation of the specific role played by Factor Xa in coagulation.     

Hexagonal crystal structure of the A-DNA octamer d(GTGTACAC) and its comparison with the tetragonal structure: correlated variations in helical parameters

The alternating DNA octamer d(GTGTACAC) has been grown in a novel hexagonal crystal form. The structure has been determined and refined to a 2-A resolution, with 51 water molecules. The A-DNA conformation is a variant of that observed for the tetragonal form of the same sequence (Jain et al., 1989) containing a bound spermine. The crystals belong to the space group P6(1)22, a = b = 32.40 A and c = 79.25 A, with one strand in the asymmetric unit. The new hexagonal structure was solved by rotation and translation searches in direct space and refined to a final R value of 12.7% by using 1561 unique reflections greater than 1.5 sigma (I). The electron density clearly shows that the penultimate A7 sugar had flipped into the alternative C2'-endo pucker. This dent in the molecule can be attributed to close intermolecular contacts. In contrast, in the tetragonal structure, the DNA is distorted in the central TA step, where the A5 backbone bonds C4'-C5' and O5'-P assume trans conformations. The hexagonal double helix more closely resembles the fiber diffraction A-DNA, compared to the tetragonal form. For instance, the tilt angle is higher (16 degrees vs 10 degrees), which is correlated with a larger displacement from the helix axis (3.5 vs 3.3), a lower rise per residue (2.9 vs 3.2), and a smaller major-groove width (6.1 vs 8.7), thus indicating that the variations in these global helical parameters are correlated. The propeller twist angles in both forms are higher for the G-C base pairs (15.3 degrees, 12.14 degrees) than for the A-T base pairs (10.8 degrees, 9.1 degrees), which is the reverse of the expected order. Unlike the tetragonal structure, the hexagonal crystal structure interestingly does not contain a bound spermine molecule. Our analysis reveals that the conformational differences between the tetragonal and hexagonal forms are not entirely due to the spermine binding, and crystal packing seems to play an important role.     

Ecoengineering high rate anaerobic digestion systems: analysis of improved syntrophic biomethanation catalysts

High performance biomethanation granules with operational specific COD removal rates of 7 kg COD removed/kg SS/d were obtained by ecoengineering conventional, granular, UASB digester sludge using a designed protocol of starvation and selection on a defined volatile fatty acid (VFA) based mineral medium. Addition of low (0.15 mM) sulfate levels to this VFA medium increased the maximum shock-load COD removal rate of the ecoengineered biomethanation granules to 9 kg COD/kg SS/d with specific acetate, propionate, and butyrate removal rates of 111, 28, and 64 mol/g SS/d. Addition of moderate (26 mM) calcium levels inhibited growth and altered the structure of granules. The general cellular, growth, stability, and performance features of these ecoengineered granules are described and discussed in relation to their use as improved biomethanation starter cultures.