Isolation and identification of β 1 → 2-d-glucans in extracellular polysaccharides of Rhizobium meliloti strain L5-30

Abstract Exopolysaccharides have been found as bound to R. meliloti cells (strain L5-30) or free in the culture medium; they consist of acidic and neutral polysaccharides. Bound and soluble neutral polysaccharides were identified as β 1 → 2 linked glucans with an average M _r value of 4000 to 6000.     

Studies on the adenosine 3′,5′-monophosphate-dependent protein kinase of Blastocladiella emersonii

Using a combination of Chromatographic and sucrose density gradient techniques under carefully controlled conditions of pH and protease inhibitors, we demonstrate that there is only one form of adenosine 3′,5′-monophosphate-dependent protein kinase in the cytosol fraction of the Blastocladiella emersonii zoospore. If any of these conditions are omitted during extract preparation, one obtains what are apparently multiple forms of the enzyme, which are in reality artifacts due to extensive endogenous proteolytic activity. This endogenous protease is stimulated by alkaline pH and inhibited by antipain. The zoospore protein kinase is similar to type II protein kinase from mammalian cells in several aspects including Chromatographic behavior on DEAE-cellulose column, conditions for subunit dissociation and reassociation, as well as the molecular weight value of the regulatory subunit.     

Veratridine-stimulated central synapses in culture: A quantitative ultrastructural analysis

Synapses in explant cultures of fetal rat neocortex at day 18 in vitro were stimulated by veratridine (10^?4M) for 20 min. The cultures were subsequently processed for electron microscopy and the synapses were analyzed by quantitative techniques, incorporating set mathematical treatment. The mean values of area, perimeter, and form factor of the presynaptic elements significantly increased following veratridine stimulation, compared to the values of control synapses. The length of the postsynaptic thickening also increased, while synaptic curvature did not change significantly in the veratridine group. A fivefold reduction was observed in the mean number of synaptic vesicles per presynaptic element and in the vesicle-terminal area ratio, following veratridine stimulation. The cytoplasm-terminal area ratio and the occurrence of vacuoles/cisternae significantly increased after veratridine application. Planar measurement of membranes (boundary length) of different presynaptic organelles revealed that the total membrane did not change significantly in the veratridine group. The data indicated an increase in volume and swelling of the pre- and postsynaptic elements, considerable depletion of synaptic vesicles, and preservation of the total presynaptic membrane following veratridine stimulation in nerve tissue culture.     

南水北调与生态平衡

水是人类赖以生存繁衍的重要资源。我国人口逐年增长,而淡水资源数量有限。多变的自然降水及频繁的旱涝灾害,一直决定着我国农业收成乃至整个国民经济的盛衰。     

DM891129菌株对沙土鼠高胆固醇血症的影响

本项研究的目的是通过动物实验来观察由健康人分离出的肠球菌DM891129菌株的降血脂作用。本实验选42只沙土鼠(gerible),雌雄各半,随机分为3组,第1组13只为正常组;第2组13只,在正常饮食中 5％胆固醇饲喂1周;第3组16只,在第2组饮食的基础上,每天以肠球菌(DM891129)菌悬液(10~9个细菌/ml)0.5ml/次,Bid灌胃一周。3组沙土鼠以眼球采血,分别检测血清胆固醇、甘油三酯和胆酸水平及粪便中肠球菌和肠杆菌水平。结果表明:第2组胆固醇水平较第1组明显升高(P<0.001)。第3组胆固醇水平较第2组明显降低(P<0.001)。相应的、第3组鼠粪便肠球菌水平明显高于第1、2组(P<0.05),而1、2组之间无显著性差异,第2、3组肠杆菌水平明显高于第1组(P<0.02、P<0.05),2、3组肠杆菌水平无显著性差异。胆酸、甘油三酯水平3组之间无显著性差异。可见DM891129菌株对沙土鼠高胆固醇饮食所致的高胆固醇血症具有明显的降低作用而对血甘油三酯、胆酸水平无显著性影响。现在对DM891129菌株体内降血胆固醇水平的机制尚不清楚,有待于进一步研究。     

Probing protein structure by solvent perturbation of nuclear magnetic resonance spectra. Nuclear magnetic resonance spectral editing and topological mapping in proteins by paramagnetic relaxation filtering.

Soluble spin labels, which "bleach" the surface proton resonances of a protein to n.m.r. measurements, can provide useful information about protein conformation and dynamics. The use of the soluble nitroxide, TEMPOL, has been explored to show the correlation of the paramagnetic perturbations of protein two-dimensional n.m.r. data with proton exposure to the free radical in hen egg-white lysozyme. The results demonstrate that the nitroxide approaches the protein randomly, and that the extent of the observed paramagnetic effects reflects the native folding pattern of the protein. A correlation of spectral simplification with the known tertiary structure establishes the feasibility of new strategies for topological mapping of surface and buried protons of the protein. Application to the elucidation of protein structure and to the study of dynamical processes is discussed.     

Extracellular Concentrations of Dopamine and Metabolites in the Rat Caudate After Oral Administration of a Novel Catechol-O-Methyltransferase Inhibitor Ro 40–7592

The effect of the systemic administration of a novel, orally active, catechol-O-methyltransferase (COMT) inhibitor, Ro 40-7592, on the in vivo extracellular concentrations of dopamine (DA) and its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), was studied by transcerebral microdialysis in the dorsal caudate of freely moving rats. Ro 40-7592 (at doses of 3.0, 7.5, and 30 mg/kg p.o.) elicited a marked and long-lasting reduction of HVA, and at doses of 7.5 and 30 mg/kg, an increase of DOPAC output, but it failed to increase DA output. The administration of L-beta-3,4-dihydroxyphenylalanine (L-DOPA, 20 and 50 mg/kg p.o.) with a DOPA decarboxylase inhibitor (benserazide) increased both HVA and DOPAC output, but failed to modify significantly extracellular DA concentrations in dialysates; in contrast, combined administration of L-DOPA+benserazide with Ro 40-7592 (30 mg/kg p.o.) resulted in a significant increase in DA output. Ro 40-7592 prevented the L-DOPA-induced increase in HVA output and markedly potentiated the increase in DOPAC output. To investigate to what extent the increase in extracellular DA concentrations was related to an exocitotic release, tetrodotoxin (TTX) sensitivity was tested. Addition of TTX to Ringer, although abolishing DA output in the absence of L-DOPA, partially reduced it in the presence of L-DOPA+Ro 40-7592 and even more so after L-DOPA without the COMT inhibitor. The results of the present study suggest that metabolism through COMT regulates extracellular concentrations of DA formed from exogenously administered L-DOPA but not of endogenous DA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunoaffinity purification and partial amino acid sequence analysis of catechol-O-methyltransferase from pig liver

Monoclonal antibodies (mAbs) against the soluble form (S-COMT) of catechol-O-methyltransferase (COMT, EC 2.1.1.6) were produced using a purified preparation of the enzyme from pig liver as antigen. The selected monoclonal antibodies recognized the enzyme with different capacities. One of them (Co60-1B/7) showed a significant cross reaction with S-COMT from rat and human liver. A protein band of 23 kDa was recognized by the mAbs on Western blots of the soluble fraction of pig liver. The mAbs were also able to recognize the membrane-bound form of the enzyme, which was found to be mainly localized in the microsomal fraction of pig and rat liver as well as of the human hepatoma cell line Hep G2. The protein bands detected in microsomes had a molecular mass of 26 kDa in pig and rat liver and displayed a slightly higher molecular mass (29 kDa) in the Hep G2 cell line. A single step method for the immunoaffinity purification of pig liver S-COMT was developed by using a Sepharose 4B column to which the mAb Co54-5F/8 was covalently coupled. Acid elution conditions were optimized to obtain the enzyme in active form with a good yield. SDS-PAGE analysis of the purified preparation revealed a single protein band with a molecular mass of 23 kDa with 154-fold enrichment in enzyme activity over the starting material. Since the N-terminus was blocked, purified enzyme preparations were cleaved with trypsin. Two fragments of 22 and 33 amino acids in length could be sequenced by Edman degradation.     

Solution conformation of salmon calcitonin in sodium dodecyl sulfate micelles as determined by two-dimensional NMR and distance geometry calculations

The 32 amino acid hormone salmon calcitonin was studied at pH 3.7 and 7.4 by two-dimensional NMR in sodium dodecyl sulfate (SDS) micelles at 310 K. The spectrum was fully assigned, and the secondary structure was obtained from nuclear Overhauser enhancement spectroscopy (NOESY), 3JHN alpha coupling constants, and slowly exchanging amide data. Three-dimensional structures consistent with NMR data were generated by using distance geometry calculations. A set of 260 interproton distances, derived from NOESY, and hydrogen-bond constraints, obtained from analysis of the amide exchange, were used. From the initial random conformations, 13 distance geometry structures with minimal violations were selected for further refinement with restrained energy minimization. In SDS, at both pHs, the main conformational feature of the hormone is an alpha-helix from Thr6 through Tyr22, thus including the amphipathic 8-22 segment and two residues of the Cys1-Cys7 N-terminal loop. The C-terminal decapeptide forms a loop folded back toward the helix. The biological significance of this conformation is discussed.