The bulk of ATP is associated to proteins in the living cell: a release kinetics study

The analysis of the nonionic detergent induced release kinetics of intracellular components is a promising approach to obtain information on the possible molecular interactions inside the living cell. The molecular interactions of ATP were studied by monitoring the nonionic detergent-induced release kinetics of ATP from cultured human cells. The observed rapid mobilization rate in the presence of Triton X 100 and the retarded release in the presence of Brij 58 were in good agreement with the detergent-induced release kinetics of proteins and K+ reported earlier. These data are consistent with the idea that the bulk of ATP is associated to the protein macromolecules of the living cell.     

Neutron irradiation of late mouse embryos (15-19 days) in utero

Pregnant female C57B1/6 mice were irradiated with a single whole-body dose of 0.5 Gy neutrons. The F1 hybrid embryos were exposed to the neutrons in utero on Day 17 +/- 2 of gestation. 178/439 (40.6%) of the irradiated fetuses and 26/217 (12%) of the control mice died within 2 weeks after birth. In both irradiated and control mice, most deaths (95 and 77%, respectively) occurred within 3 days of birth: most animals in both groups died on Day 2. There was no significant difference in the number of living young born per litter (7.2) between the neutron-irradiated mothers and their unirradiated controls. The irradiated mice weighed significantly less than their controls. On the first day after birth, body weights of mice irradiated in utero averaged only 85% of control weights. Body weights did not reach control levels until 6 months after birth. Several organs were weighed at regular intervals in both irradiated and control mice. Spleens and thymus glands showed no significant differences between the two groups. The livers and kidneys of the irradiated mice weighed slightly less than their controls. The brain weight of 21-day-old neutron-irradiated mice was 30-35% less than control brains. The weight loss of the brain was not only a relative loss, but also an absolute one, based on brain weight/body weight ratios. Histological analysis of the central nervous system showed pycnotic nuclei, inhibition of mitosis in neuroblasts, and cell death in the irradiated brains. The weight reduction of the brain was not due to water loss. Our hypothesis is that the early mortality after birth is related to the killing of the radiation-sensitive neuroblasts. When newborn mice (1-7 days old) were irradiated in vivo with the same neutron dose of 0.5 Gy, neither the reduction in brain weight nor the early mortality was observed. The early deaths of the neutron-irradiated mouse embryos does not appear to be caused by either the hematological or the gastrointestinal radiation syndrome.     

A comparison of liquid hot water and steam pretreatments of sugar cane bagasse for bioconversion to ethanol.

Sugar cane bagasse was pretreated with either liquid hot water (LHW) or steam using the same 25 l reactor. Solids concentration ranged from 1% to 8% for LHW pretreatment and was > or = 50% for steam pretreatment. Reaction temperature and time ranged from 170 to 230 degrees C and 1 to 46 min, respectively. Key performance metrics included fiber reactivity, xylan recovery, and the extent to which pretreatment hydrolyzate inhibited glucose fermentation. In four cases, LHW pretreatment achieved > or = 80% conversion by simultaneous saccharification and fermentation (SSF). > or = 80% xylan recovery, and no hydrolyzate inhibition of glucose fermentation yield. Combined effectiveness was not as good for steam pretreatment due to low xylan recovery. SSF conversion increased and xylan recovery decreased as xylan dissolution increased for both modes. SSF conversion, xylan dissolution. hydrolyzate furfural concentration, and hydrolyzate inhibition increased, while xylan recovery and hydrolyzate pH decreased, as a function of increasing LHW pretreatment solids concentration (1-8%). These results are consistent with the notion that autohydrolysis plays an important. if not exclusive, role in batch hydrothermal pretreatment. Achieving concurrently high (greater than 90%) SSF conversion and xylan recovery will likely require a modified reactor configuration (e.g. continuous percolation or base addition) that better preserves dissolved xylan.     

Technical considerations in the gas chromatographic analysis of desipramine

Procedures are presented for minimizing variability in and interferences with the gas chromatographic determination of desipramine in plasma. Careful consideration of procedures for sample collection and storage, drug separation from matrix components, and chromatography appears to be a prerequisite for avoiding inaccurate and imprecise determinations of this antidepressant, especially at levels below 20 μg/l. Numerous pitfalls are examined and optimal conditions for obviating them are presented.     

Molten globule-like state of cytochrome c induced by polyanion poly(vinylsulfate) in slightly acidic pH.

The effect of polyanion, poly(vinylsulfate), used as a model of negatively charged surface, on ferric cytochrome c (ferricyt c) structure in acidic pH has been studied by absorbance spectroscopy, circular dichroism (CD), tryptophan (Trp) fluorescence and microcalorimetry. The polyanion induced only small changes in the native structure of the protein at neutral pH, but it profoundly shifted the acid induced high spin state of the heme in the active center of cyt c to a more neutral pH region. Cooperativity of the acidic transition of ferricyt c in the presence of the polyanion was disturbed, in comparison with uncomplexed protein, as followed from different apparent pK(a) values observed in a distinct regions of the ferricyt c electronic absorbance spectrum (4.55+/-0.08 in the 620 nm band region and 5.47+/-0.15 in the Soret region). The ferricyt c structure in the complex with the polyanion at acidic pH (below pH 5.0) has properties of a molten globule-like state. Its tertiary structure is strongly disturbed according to CD and microcalorimetry measurements; however, its secondary structure, from CD, is still native-like and ferricyt c is in a compact state as evidenced by quenched Trp fluorescence. These findings are discussed in the context of the molten globule state of proteins induced on a negatively charged membrane surface under physiological conditions.     

Spectrophotometric detection of the interaction between cytochrome c and heparin.

Heparin inhibits transport of electrons from reduced cytochrome c to cytochrome c oxidase. The effect is due to the interaction of heparin with cytochrome c. It has been observed that binding of heparin to the reduced or oxidized cytochrome c changes the spectrum of cytochrome c at the Soret region. Affinity chromatography of heparin in cytochrome c immobilized to thiol-Sepharose shows that commercial heparin is eluted in the low-affinity and high-affinity fractions. Both participate in the interaction with cytochrome c. Polylysine induces decay of the cytochrome c-heparin complex.     

Prevention of thermal induced aggregation of cytochrome c at isoelectric pH values by polyanions

Differential scanning calorimetry, viscometry, optical and CD spectroscopy were used to characterize the influence of two polyanions, poly(vinylsulfate) (PVS), and poly(4-styrene-sulfonate) (PSS) on thermal transition reversibility of ferricytochrome c at or near isoelectric pH. In these conditions, both PVS and PSS enhance the thermal transition reversibility of cytochrome c by preventing the aggregation of denatured protein molecules. Data indicate that the polyanions are in complex with cytochrome c that is stabilized by synergistic effect of Coulombic and non-Coulombic interactions.