Glutamine Synthetase of the Human Brain: Purification and Characterization

Glutamine synthetase (GS) isolated from human brain formed a single band on sodium dodecyl sulfate-polyacrylamide gel with a molecular weight of 44,000. The enzyme had a specific activity of 179.2 U/mg protein when assayed by measuring the rate of the formation of gamma-glutamylhydroxamate using hydroxylamine as a substrate. In the presence of manganese ions, the relative activity of human brain GS was much lower than that of the sheep brain enzyme. The suppression of activity by increasing the ADP concentration, however, was less marked in the human enzyme than that in the sheep enzyme. Antibodies were raised in rabbits against the purified enzyme. The double-immunodiffusion technique disclosed cross-reactivities among GSs isolated from human, sheep, and rat brains, but the enzymes were not immunologically identical. Immunohistochemically, GS was localized in the cytoplasm of astrocytes in the human and rat brains and in pericentral hepatocytes of the liver.     

The different effects of recombinant human tumor necrosis factor on rat fibrosarcoma sublines

Summary The antitumor effect of recombinant human tumor necrosis factor (rH-TNF) on two clones of rat fibrosarcoma with different metastatic potential to lymph nodes was examined. The colony formation of clone A, which has high metastatic potential, was completely inhibited by continuous exposure to rH-TNF at 50 U/ml. In contrast, colony formation of clone G, which has low metastatic potential, was not inhibited by high concentrations of rH-TNF (10,000 U/ml). The inhibitory effect of rH-TNF on colony formation by clone A was also observed with a 1-h exposure to rH-TNF. This effect was time and concentration dependent, as determined by the colony assay, ³H-thymidine uptake assay, and ⁵¹Cr-release assay. ³H-thymidine and ³H-uridine uptake per cell of clone A exposed to rH-TNF was not decreased. This suggests that the mechanisms of the antitumor effect of rH-TNF were not due to inhibition of DNA and RNA synthesis of tumor cells. In vivo growth and lymph node metastases of clone A inoculated i.p. to Donryu strain rats were completely suppressed by 14 consecutive i.p. injections of 10⁵ or 10⁶ U/kg per day of rH-TNF. On the other hand the growth of clone G was not influenced by rH-TNF administration.     

Application of an enzyme-linked immunosorbent assay for screening of T-2 toxin-producing Fusarium spp.

Culture filtrates of Fusarium species were subjected without clean-up procedures to an indirect competitive enzyme-linked immunosorbent assay with anti-T-2 toxin monoclonal antibody. Fusarium sporotrichioides, F. poae, F. tricinctum, and F. culmorum strains were positive for T-2 toxin, with a minimum detection limit of 5 pg per assay (100 pg/ml of culture filtrate), and the assay data correlated well with the gas-liquid chromatographic data.     

Specific receptors for thromboxane A2 in cultured vascular smooth muscle cells of rat aorta

The specific binding site for thromboxane A2 (TXA2) was studied in cultured vascular smooth muscle cells (VSMC) of the rat aorta. [3H]SQ29,548, a potent and selective TXA2 receptor antagonist, displayed high-affinity and specificity, as well as saturable and displaceable binding to rat VSMC in culture. Scatchard analysis of equilibrium binding at 24 degrees C revealed a single class of binding sites with a Kd of 1.7 nM and a Bmax of 8.0 fmol/10(6) cells. A series of TXA2 receptor antagonists completely suppressed [3H]SQ29,548 binding to rat VSMC, and the rank order of their inhibitory potencies (Ki) correlated well with the potencies for suppression of the U46619-induced contraction of rat thoracic aorta. These results suggest that specific binding sites for [3H]SQ29,548 represent the TXA2 receptor in rat VSMC.     

Branching Photocycle of Sensory Rhodopsin in Halobacterium Halobium

Temperature effect of the photocyle of sensory rhodopsin (sR) was studied by nanosecond spectroscopy. Though the formation yield of sR_M (sR₃₇₀) was sharply decreased with temperature, those of sR_K (sR₆₈₀) and sR_L were insensitive to temperature changes. These results show the existence of the branching process back to sR from sR_L. The absorption maxima for sR_K and sR_L were 595 ± 5 and 555 ± 15 nm, respectively.     

A New Myelin-Deficient Mutant Hamster: Biochemical and Morphological Studies

Biochemical and morphological studies were done on a new trembling mutant hamster CBB. The yield of myelin from the mutant was 30 and 40% of the control at 46 and 140 days of age, respectively, but myelin composition and 2',3'-cyclic nucleotide-3'-phosphohydrolase (CNPase) activity were normal. Morphologically, about 18% of the axons were myelinated in the mutant optic nerve at 46 days of age, in which the myelinated fibers were those with larger diameters (more than 0.6 micron), while the control had a peak at 0.4 micron in diameter. The ultrastructure and thickness of compact myelin lamellae in the mutant were normal. Myelination and the structure of peripheral nerve myelin appeared normal. The results indicate that the essential defect is the delay and arrest of myelination in the CNS, which is probably caused by either a decreased rate of synthesis of myelin components in oligodendrocytes or a defect in the oligodendrocyte-axon recognition in smaller axons.     

A versatile bacterial enzyme: l-methionine γ-lyase

The properties and application of l-methionine γ-lyase [methioninase, l-methionine methanethiol-lyase (deaminating), EC 4.4.1.11], a pyridoxal 5′-phosphate enzyme, purified from Pseudomonas putida and Aeromonas sp. are presented. The enzyme has multicatalytic functions: it catalyses α,γ-elimination and γ-replacement reactions of l-methionine and its analogues (e.g. ethionine, homocysteine, O-acetylhomoserine and selenomethionine), α,β-elimination and β-replacement reactions of l-cysteine and its analogues (e.g. S-methylcysteine, O-acetylserine and Se-methylselenocysteine), deamination and γ-addition of vinylglycine, and deuterium labelling at the α and β positions of l-methionine and other straight-chain l-amino acids. These reactions are applicable to the synthesis of various optically active sulphur and selenium amino acids, preparation of deuterium or tritium labelled l-amino acids, and determination of sulphur amino acids. In addition, the enzyme shows potent anti-neoplastic activity.     

An investigation for the occurrence of C4 photosynthesis in the Cyperaceae from Australia

Two hundred and twenty species of 38 genera in the Cyperaceae from Australia were examined for the possible occurrence of the C₄ photosynthesis and the anatomical features of leaves and culms. The Kranz type of anatomy and the carbon isotope ratios typical of C₄ plants were found in 84 species in the following six genera of four tribes belonging to subfamily Cyperoideae:Bulbostylis, Crosslandia, andFimbristylis (Fimbristylideae);Lipocarpha (Lipocarpheae);Cyperus (Cypereae);Rhynchospora (Rhynchosporeae). The anatomical observation revealed that the C₄ species possessed any one of the three Kranz anatomical types found by previous investigators. It was suggested that in the Cyperaceae the C₄ syndrome evolved independently within several taxa of the subfamily. The relative distribution of C₃ and C₄ species of the Cyperaceae in Australia was investigated by use of floristic data. It was recognized that the C₄ species dominated in the northern part of the continent which was characterized by tropical and subtropical savannas and hot dry areas with summer rainfall, and the C₃ species in the southern part, which contained temperate areas and mediterranean climatic areas with winter rainfall.     

Variations of Natural 13C Abundances in Leguminous Plants

The natural ¹³C abundance (¹³C value) of the field-grown leguminousplants (soybean, kidney bean, pea, azuki bean, mung bean, peanutand cowpea) was investigated by mass spectrometry with a precisionbetter than %0.2 for ¹³C. Among organs of premature plants,the leaves had the most negative values, and the nodules generallyhad the least negative values, and other organs, fruits, stemsand roots, showed intermediate values. In the soybeans so farinvestigated, the grains of nodulating plants exhibited higher¹³C values than nonnodulating lines. The ¹³C values of the grainsvaried depending on the species: peanuts showed the most negativevalues. Possible causes underlying these variations are discussed. ³Institute of Biological Sciences, University of Tsukuba, Sakura-mura,Ibaraki 305, Japan. (Received December 3, 1982; Accepted May 25, 1983)