Rat liver cysteine sulfinate decarboxylase: purification, new appraisal of the molecular weight and determination of catalytic properties.

Rat liver cystein sulfinate decarboxylase (L-cystein sulfinate carboxylase) was purified approximately 500-fold. By cellulose acetate and polyacrylamide gel electrophoresis or by analytical ultracentrifugation, the purified enzyme appears to be nearly homogeneous. The Stokes radius (3.4 nm) and sedimentation coefficient (6.5 S) were determined. The molecular weight, calculated and experimentally estimated is around 100 000 and the enzyme is constituted of two identical subunits whose molecular weights are 55 000. The role of pyridoxal phosphate as coenzyme was demonstrated and the requirement for free sulhydryl groups for activity was studied. The ability of native pure cysteine sulfinate decarboxylase to also decarboxylate cysteate was stressed: therefore, we concluded that in rat liver a single protein catalyzed both reactions, although only the decarboxylation of cysteine sulfinate is of physiological interest.     

Circadian variations of the effects of ethanol and of blood ethanol values in the healthy adult man. Chronopharmacological study]

Six healthy young men (22 to 26 years) who had fasted for 12 hours volunteered for this study (subject synchronization: diurnal activity from 07(00) to midnight and nocturnal rest). A set dose of ethanol (0.67 g/kg body weight) was ingested at the fixed (and random) hours of 07(00), 11(00), 19(00) and 23(00), with a week between tests. A set of physiological variables: psychological tests (selft-rating of mood, of physical vigor and of ebriety, tempo, random number addition test); physical variables (heart rate, systolic and diastolic blood pressure, peak expiratory flow, oral temperature and grip strength); blood variables (plasma ethanol, cortisol, lactic acid, pyruvic acid, glucose and erythrocyte K+) and urinary variables (volume, epinephrine, nor-epinephrine and 5-HIAA) were documented at least at 4 hourly intervals and set times. The cosinor method was used for chronobiological statistical analyses. The parameters characterizing the ethanol pharmacokinetics (chronopharmacokinetics) demonstrated a circadian rhythm (p less than 0.05): e. g. the peak height of ethanolemia is greater when ethanol is ingested at 07(00) than at other times. Also a circadian rhythm in biosystems susceptibility can be demonstrated (p less than 0.05) (chronesthesy) with a peak time not necessarily corresponding either to that of ethanolemia or to that of other variables. The overall circadian changes in ethanol effects (chronergy) can be viewed as a combination of both ethanol chronesthesy and chronokinetics.     

Makeup of free intracellular amino acids in Cunninghamella elegans growing on media with hydrocarbons]

The rate of growth of Cunninghamella elegans (--) 1204 is higher on a mineral medium with glucose (6.56 g/litre) than on a mineral medium containing undecane, tridecane, and pentadecane (0.72--0.87 g/litre); all glutamic acid is consumed only from the medium with glucose. The cells contain 15--16 free amino acids and 1--2 amides, glutamic and aspartic acids and alanine prevailing. The culture grown on the medium with glucose contains asparagine, and the cells cultivated on the medium with alkanes contain histidine. Non-proteinogenous aminobutyric acids were found in the pool of the cells grown on all tested media with an exception of the medium containing undecane.     

Effect of a "direct" hemolytic factor and phospholipse A2 from Middle-Asian cobra venom on erythrocytes]

The effect of "direct" hemolytic factor (DHF) and phospholipase A2, isolated and purifed from Middle-Asian cobra venom on human erythrocytes is studied. DHF is found to increase the hemolytic effect of phospholipase A2 Ca2+ ions inhibit the hemolytic effect of DHF, but they increase the hemolysis in the presence of DHF and phospholipse A2. Heparin re moves the increasing effect of DHF. Possible mechanisms of the hemolytic effect of DHF and phospholipase A2 are discussed.     

Thermal stability of electron transport in PS II membranes and particles from the thermophilic cyanobacteria

Thermal stability of the ferricyanide (FC) and dichlorophenolindophenol (DCIP) reducing reactions was investigated in isolated membrane preparations and PS II particles with active water splitting system from the cyanobacterium Synechococcus elongatus. In a hypotonic medium, the thermostability was seen to be much higher for the DCIP than for the FC reduction reaction. After the addition of high concentrations of polyethylene glycol (Mwt = 4000) or sodium citrate to the medium, the FC reduction reaction appeared to be more temperature resistant. Data on the effects of temperature, DCMU and detergents on the electron transfer rate in PS II provide evidence suggesting that the different thermal stabilities of the two reactions are due to different physico-chemical properties of the electron donor sites to FC and DCIP. The data suggest that regions of contact between individual macromolecular complexes of the electron transport chain are the most labile sites of the photosynthetic apparatus. The role of the composition and properties of the intracellular medium on thermostability are emphasized.     

Efficient colonization of the endophytes Herbaspirillum huttiense RCA24 and Enterobacter cloacae RCA25 influences the physiological parameters of Oryza sativa L. cv. Baldo rice

Several important bacterial characteristics, such as biological nitrogen fixation, phosphate solubilization, 1-aminocyclopropane-1-carboxylate deaminase activity and production of siderophores and phytohormones, can be assessed as plant growth promotion traits. Our aim was to evaluate the effects of nitrogen fixing and indole-3-acetic acid (IAA) producing endophytes in two Oryza sativa cultivars (Baldo and Vialone Nano). Three bacteria, Herbaspirillum huttiense RCA24, Enterobacter asburiae RCA23 and Staphylococcus sp. 377, producing different IAA levels, were tested for their ability to enhance nifH gene expression and nitrogenase activity in Enterobacter cloacae RCA25. Results showed that H. huttiense RCA24 performed best. Improvement in nitrogen fixation and changes in physiological parameters such as chlorophyll, nitrogen content and shoot dry weight were observed for plants co-inoculated with strains RCA25 and RCA24 in a 10:1 ratio. Based on confocal laser scanning microscopy analysis, strain RCA24 was the best colonizer of the root interior and the only IAA producer located in the same root niche occupied by RCA25 cells. This work shows that the choice of a bio-inoculum having the right composition is one of the key aspects to be considered for the inoculation of a specific host plant cultivar with microbial consortia.     

The quaternary structure of bovine heart mitochondrial creatine kinase

Crosslinking of subunits of the high molecular weight oligomer of bovine heart mitochondrial creatine kinase (CKm) by dimethyl suberimidate and subsequent electrophoresis in the presence of sodium dodecyl sulfate gives eight protein bands. An increase in the time course of the enzyme crosslinking reaction results in the protein accumulation in the high molecular weight bands. Evidence has been obtained suggesting that crosslinking involves only the intraoligomeric contact areas. It is concluded that bovine heart CKm is an octamer. Crosslinking of intersubunit contacts in the octameric form of the enzyme by various diimidates has been carried out. The data obtained suggest that within the octamer the CKm subunits have a quasispherical rather than planar arrangement. This finding is supported by electron microscopy data.     

Differential Association of the Conserved SUMO Ligase Zip3 with Meiotic Double-Strand Break Sites Reveals Regional Variations in the Outcome of Meiotic Recombination

During the first meiotic prophase, programmed DNA double-strand breaks (DSBs) are distributed non randomly at hotspots along chromosomes, to initiate recombination. In all organisms, more DSBs are formed than crossovers (CO), the repair product that creates a physical link between homologs and allows their correct segregation. It is not known whether all DSB hotspots are also CO hotspots or if the CO/DSB ratio varies with the chromosomal location. Here, we investigated the variations in the CO/DSB ratio by mapping genome-wide the binding sites of the Zip3 protein during budding yeast meiosis. We show that Zip3 associates with DSB sites that are engaged in repair by CO, and Zip3 enrichment at DSBs reflects the DSB tendency to be repaired by CO. Moreover, the relative amount of Zip3 per DSB varies with the chromosomal location, and specific chromosomal features are associated with high or low Zip3 per DSB. This work shows that DSB hotspots are not necessarily CO hotspots and suggests that different categories of DSB sites may fulfill different functions.     

Impact of Anesthetics on Immune Functions in a Rat Model of Vagus Nerve Stimulation

Vagus nerve stimulation (VNS) has been successfully performed in animals for the treatment of different experimental models of inflammation. The anti-inflammatory effect of VNS involves the release of acetylcholine by vagus nerve efferent fibers inhibiting pro-inflammatory cytokines (e.g. TNF-α) produced by macrophages. Moreover, it has recently been demonstrated that splenic lymphocytic populations may also be involved. As anesthetics can modulate the inflammatory response, the current study evaluated the effect of two different anesthetics, isoflurane and pentobarbital, on splenic cellular and molecular parameters in a VNS rat model. Spleens were collected for the characterization of lymphocytes sub-populations by flow cytometry and quantification of cytokines secretion after in vitro activation. Different results were observed depending on the anesthetic used. The use of isoflurane displayed a non-specific effect of VNS characterized by a decrease of most splenic lymphocytes sub-populations studied, and also led to a significantly lower TNF-α secretion by splenocytes. However, the use of pentobarbital brought to light immune modifications in non-stimulated animals that were not observed with isoflurane, and also revealed a specific effect of VNS, notably at the level of T lymphocytes’ activation. These differences between the two anesthetics could be related to the anti-inflammatory properties of isoflurane. In conclusion, pentobarbital is more adapted than isoflurane in the study of the anti-inflammatory effect of VNS on an anesthetized rat model in that it allows more accurate monitoring of subtle immunomodulatory processes.