Correlation between Possession of a Respiration-Dependent Na Pump and Na Requirement for Growth of Marine Bacteria

The possession of a respiration-dependent primary sodium pump and the requirement of Na for growth were investigated in bacterial isolates from marine environments. The bacteria in which NADH oxidase specifically required Na for maximum activity were believed to possess a primary sodium pump. All bacteria that failed to grow without the addition of NaCl possessed a primary Na pump. All bacteria that had no primary Na pump grew without additional NaCl. The primary Na pump seems to be involved in the Na requirement of marine bacteria, and this can be regarded as a criterion for the definition of marine bacteria.     

BsoAI--a new site specific endonuclease from Bacillus coagulans

A new site-specific endonuclease was detected in toluene lysates of Bacillus coagulans AUCM B-732 and designated as BcoAI. The enzyme was purified by fractionation of the cell-free extract in the two-phase PEG/dextran system followed by chromatography on DEAE-sepharose and phosphocellulose and shown to be free of nonspecific nucleases and phosphatases. BcoAI has three cleavage sites on lambda DNA, but does not cleave SV40, pBR322 and pUC19 DNA. BcoAI recognizes the sequence 5' CAC decreases GTG 3' on double-stranded DNA and cleaves it as indicated by the arrow to yield blunt-ended DNA fragments. Thus, BcoAI is a true isoschizomer of PmaCI from Pseudomonas maltophila C.     

Determination of the spatial structure of insectotoxin 15A from Buthus erpeus by (1)H-NMR spectroscopy data]

The solution structure of insectotoxin 15A (35 residues) from scorpion Buthus eupeus was determined on the basis of 386 interproton distance restraints 12 hydrogen-bonding restraints and 113 dihedral angle restraints derived from 1H NMR experiments. A group of 20 structures was calculated with the distance geometry program DIANA followed by the restrained energy minimization with the program CHARMM. The atomic RMS distribution about the mean coordinate position is 0.64 +/- 0.11 A for the backbone atoms and 1.35 +/- 0.20 A for all atoms. The structure contains an alpha-helix (residues 10-20) and a three-stranded antiparallel beta-sheet (residues 2-5, 24-28 and 29-33). A pairing of the eight cysteine residues of insectotoxin 15A was established basing on NMR data. Three disulfide bridges (residues 2-19, 16-31 and 20-33) connect the alpha-helix with the beta-sheet, and the fourth one (5-26) joins beta-strands together. The spatial fold of secondary structure elements (the alpha-helix and the beta-sheet) of the insectotoxin 15A is very similar to those of the other short and long scorpion toxins in spite of a low (about 20%) sequence homology.     

Enzymatic characterization of Vibrionaceae strains isolated from environment and cold-blooded animals.

Enzymatic profiles were determined by the API ZYM system for 15 strains of non 01 Vibrio cholerae, 4 strains of V. metschnikovii, 9 strains of V. anguillarum, 6 strains of Plesiomonas shigelloides and 115 strains motile Aeromonas sp. All of the tested strains produced alkaline phosphatase, leucine aminopeptidase and did not possess alpha-fucosidase and alpha-mannosidase. Some differences in enzymatic activities among the tested Vibrionaceae strains were noted. The strains of non 01 V. cholerae, V. metschnikovii, V. anguillarum and P. shigelloides did not produce trypsin, whereas all of the tested Vibrio sp. strains appeared to be positive for this enzyme. Only the strains of P. shigelloides produced BI-Phospho-hydrolase. The lack of acid phosphatase activity was observed among the strains of V. anguillarum.     

Spatial structure of the basic chain of the neurotoxin I molecule from the venom of cobra Naja naja oxiana and its crystal packing

The structure of the alpha-carbon chain was solved by molecular replacement method at 2.7 A resolution. Neurotoxin I (NTX-I) is one of the main protein components purified from the venom of the central asian cobra Naja naja oxiana. NTX-I is known to bind specifically to acetylcholine receptors thus preventing the transmission of the neuroconductivity signal from synapses to muscles. NTX-I crystals were grown either by vapour diffusion or dialysis methods using specially prepared microdialysis cell. The intensities of reflections from native NTX-I crystals were measured in the range of 38.0-2.1 A-1 by omega-scan method with a Syntex P21 diffractometer operated in automatic regime. To determine the position and mode of packing of NTX-I molecule in unit cell program packages MERLOT and BLANC were applied running on a NORD-500 computer.     

Partitioning of beta-galactosidase fusion proteins in PEG/potassium phosphate aqueous two-phase systems.

Four different beta-galactosidase fusion proteins have been partitioned in poly(ethylene glycol) (PEG) 4000/potassium phosphate aqueous two-phase systems. The partition coefficients (K) of staphylococcal protein A-beta-galactosidase (SpA beta gal) (K = 3.5) and staphylococcal protein A-streptococcal protein G-beta-galactosidase (AG beta gal) (K = 2.8) were compared with the partition coefficients of their constituent molecules, beta-galactosidase, SpA, and protein AG. It was found that by fusing beta-galactosidase to the smaller proteins SpA and protein AG, their partition coefficients were increased four to five times. Experimental data were fitted into, and found to agree with, the Albertsson partition model of interacting molecules. The compatibility with PEG and potassium phosphate of beta-galactosidase, SpA, and two different versions of the SpA beta gal protein, displayed as precipitation curves, showed a relationship to the protein partition coefficients in PEG/potassium phosphate systems. High solubility in one phase component was accompanied by preferential partitioning to the phase rich in the same component in the PEG/potassium phosphate system. Also, a changed linker region in SpA beta gal resulted in a more soluble protein. This, together with the improved K values of the target proteins by fusion, shows that it is possible to use beta-galactosidase as an affinity handle.     

Characteristics of a novel high viscosity polysaccharide,methylan, produced byMethylobacterium organophilum

Summary A new anionic high molecular weight polysaccharide, Methylan, was produced byMethylobacterium organophilum from methanol as a sole source of carbon and energy under the specific culture conditions. By GPC and light scattering, the molecular weight was determined to be 2–4×10⁶ dalton and the distribution of molecular weight was very homogeneous. Methylan was composed of carbohydrate (80%), uronic acid (12%), protein (6%) and pyruvic acid (5%). The sugar composition of Methylan was identified as glucose, galactose and mannose with the approximate molar ratio of 232. Methylan solution showed a pseudoplastic non-Newtonian fluid property at the concentration above 0.05%. At the concentration of 1% Methylan solution, the consistency index was 18,000 centipoise which was almost 10 times higher than that of Xanthan and the flow behavior index was 0.15.     

Supplement of nutrients for effective cultivation of hepatitis B surface antigen-producing recombinant yeast

Summary In the hepatitis B surface antigen (HBsAg)-producing recombinant yeast culture medium, the supply of Bacto-yeast nitrogen base without amino acids was found to be inadequate due to the lack of the several kinds of vitamins and trace elements. When the culture medium for this recombinant yeast was supplemented with sufficient vitamins and trace elements, its growth, HBsAg production and the stability of plasmid were improved.     

Changes in levels of mRNA coding for catecholamine synthesizing enzymes and neuropeptide Y in the adrenal medulla of the newborn rat.

We have measured levels of mRNA coding for the catecholamine synthesizing enzymes tyrosine hydroxylase (TH), dopamine beta-hydroxylase (D beta H), phenylethanolamine N-methyltransferase (PNMT) and for neuropeptide Y (NPY) in rat adrenal medulla by using in situ hybridization histochemistry. Ages of one day before birth (E21), 12 h, 24 h, 2 days and 4 days after birth and in adults were studied. TH, D beta H and NPY mRNA levels increased markedly postnatally. Twelve hours after birth the levels of mRNA for TH, D beta H and NPY were, respectively, 512 +/- 18%, 370 +/- 24% and 253 +/- 21% of E21 levels. At 24 h of age NPY mRNA level was 437 +/- 73% of fetal value. In contrast, the levels of mRNA coding for PNMT increased more slowly and reached 196 +/- 9% of E21 level on postnatal day four and was further increased in adult rats.     

Interannual variability of phytoplankton productivity and related parameters in Lake Constance: no response to decreased phosphorus loading?

In meso-eutrophic Lake Constance (Germany-Austria-Switzerland),phytoplankton bioraass, pigments and water transparency, aswell as primary productivity, have been followed between 1980and 1989. During this period, municipal phosphorus loading declinedsignificantly. Since 1981, soluble reactive phosphorus (SRP)concentrations during deep lake mixing have decreased from 3.0to currently 1 6 mmol m^–3 at a rate of 7% year^–1.Nitrate concentrations, by contrast, continued to rise. Duringthe period of maximum phosphorus loading, flushing through theoutlet and sedimentation were about equally important sinksof phosphorus from the euphotic zone. Recently, however, sedimentationand subsequent burial of P in the bottom deposits contributedabout three-quarters to the overall P-losses from the systemMain reasons for this shift are unchanged settling fluxes ofphosphorus out of the euphotic zone and decreasing concentrationsof total phosphorus in the water. Only during spring, do concentrations of soluble reactive phosphoruswithin the euphotic zone decrease in proportion to the formationof particulate organic matter. Later during the season, euphoticSRP concentrations continue to be low but are no longer matchedby high plankton biomass because phosphorus is efficiently removedby settling of particles In spite of the observed dramatic decreasein phosphorus loading since 1980, chlorophyll concentrationsand water transparency, as well as annual phytoplankton productivity(300 g C m^–2), have not shown a consistent downward trend.However, the intensity of phosphorus regeneration within theeuphoric zone, which can be used as a measure of the degreeof nutrient limitation, is likely to have increased significantlyThe most probable explanation for the insensitivity of importanttrophic state indicators to reduced nutrient loading is that,in Lake Constance, biomass accumulation to a greater extentis controlled by losses, mainly grazing by zooplankton and sedimentation,than by primary resources. This is concluded from the observationthat phytoplankton biomass always falls far short of the nutrient-dependentcarrying capacity of the system.