Metastability and polymorphism in the gel phase of 1,2-dipalmitoyl-3-SN-phosphatidylcholine. A Fourier transform infrared study of the subtransition.

Fourier transform infrared spectroscopy was used to study the metastability of 1,2-dipalmitoyl-3-sn-phosphatidylcholine (DPPC) at temperatures near 0 degrees C. It was found that when DPPC is incubated at 2 degrees C for three days the two-dimensional acyl chain packing changes from one resulting in spectra typical of an orthorhombic subcell to one resembling that found in triclinically packed acyl systems. This transition proceeds in two stages. The first step, requiring less than one day, approximates first-order kinetics; the second stage proceeds with second- or higher-order kinetics. Comparison of spectra recorded at -36 degrees C with and without prior incubation at 2 degrees C shows that there are two stable low temperature forms of DPPC; that is, DPPC is metastable only within a narrow temperature range. A study of the thermotropic behavior in the range 0-45 degrees C shows that the subtransition near 15 degrees C is a transition from the alternate form to one with orthorhombic characteristics. Spectral changes at the pretransition and the main phase transition demonstrate that there are differences in behavior that are related to the thermal history of the sample.     

Temperature-dependent variation in the synthesis of streptococcal group-specific carbohydrate. II. Biosynthetic studies in group A and variant strains.

The biosynthesis of the cell wall polysaccharide and peptidoglycan of group A and A-486-Var streptococci was studied with N-acetyl-[14C]glucosamine, UDP-N-acetyl-[14C]glucosamine, and [14C]glucose. The incorporation of N-acetyl-[14C]-glucosamine into the cell wall four times greater in the A-486-Var cells than in the group A cells. However, the percentage of the total label incorporated into the cell wall polysaccharide at 37 degrees C by the A-486-Var strain was 12%, compared with 66% for the group A cells. When the A-486-Var was grown at 22 degrees C, the proportion of the label incorporated into the cell wall polysaccharide increased to 41%. At 37 degrees C, N-acetyl-[14C]glucosamine was incorporated preferentially into the peptidoglycan of the A-486-Var; almost three times as much of the label was incorporated into the peptidoglycan at 37 degrees C as was incorporated at 22 degrees C. Studies with protoplast membranes of these organisms showed similar differences, with a fourfold greater uptake of UDP-N-acetyl-[14C]glucosamine by the A-486-Var membranes at both incubation temperatures. These studies suggest that a defect in the incorporation of N-acetylglucosamine into the side chain of the polysaccharide is present in the A-486-Var strain at a step following the synthesis of UDP-N-acetylglucosamine. This defect, which may involve the UDP-N-acetylglucosamine transferase, is temperature dependent in the A-486-Var strain.     

A psychrophilic strainActinoplanes sp. 220

The strain designated Actinoplanes sp. 220 differed in its characteristics from other strains of the genus Actinoplanes listed in Bergey's Manual (1974). The strain belongs to psychrophilic culture growing within the range of 0-30 degrees C. The optimal temperature for growth on yeast--malt agar is 10-23 degrees C. Cultures transferred at 23 and 28 degrees C differed in morphological and physiological properties, enzyme activity and pigmentation in standard media. Submerged culture transferred at 28 degrees C inhibited growth of Bacillus subtilis ATCC 6633 and ATCC 9945. LL-2,6-Diaminopimelic acid was chromatographically detected in the submerged mycelium of this culture. This compound was not found in the mycelium of the original culture transferred at 23 degrees C. The cultures did not substantially differ in the composition of other amino acids contained in larger quantities in the mycelium.     

Dynamics of the phosphate group in phospholipid bilayers. A 31P nuclear relaxation time study.

The spin-lattice relaxation time of the 31P nucleus in the phosphate group of egg yolk phosphatidylcholine multilamellar dispersions has been investigated at four resonant frequencies (38.9, 81.0, 108.9, and 145.7 MHz) in the temperature range from -30 degrees to 60 degrees C. The observed frequency dependence of the relaxation indicates that both dipolar relaxation and relaxation due to anisotropic chemical shielding are significant mechanisms. The experimental data have thus been modeled assuming both mechanisms and the analysis has allowed the contribution of each to the relaxation to be determined along with the correlation time for the molecular reorientation as a function of temperature. Dipolar relaxation was found to dominate at low nuclear magnetic resonance frequencies while at high frequencies the anisotropic chemical shift dominates. The correlation time of the phosphate group is on the order of 10(-9) s at 60 degrees C and increases to approximately 10(-7) s at -30 degrees C. It is observed that the freezing of the buffer which occurs at approximately -8 degrees C has a significant effect on the phosphate group reorientation. This effect of the freezing is to change the activation energy for the phosphate group reorientation from 16.9 KJ/mol above -8 degrees C to 32.5 KJ/mol below -8 degrees C.     

Thermoregulation in the Angolan free-tailed bat Mops condylurus: A small mammal that uses hot roosts.

The Angolan free-tailed bat (Mops condylurus) uses roosts that often exceed 40 degrees C, an ambient temperature (Ta) that is lethal to many microchiropterans. We measured the physiological responses of this species at Ta's from 15 degrees to 45 degrees C. Torpor was commonly employed during the day at the lower Ta, but the bats generally remained euthermic at night, with a mean body temperature (Tb) of 35.2 degrees C. Metabolic rate reflected the pattern of Tb, increasing with falling Ta at night but decreasing during the day. Metabolic rate and evaporative losses were lower in torpid than in euthermic bats. Body temperature increased at each Ta >35 degrees C and was 43 degrees C at Ta of 45 degrees C. At Ta of 40 degrees C bats increased dry thermal conductance and evaporative heat loss compared to lower Ta. At 45 degrees C dry thermal conductance was lower than at 40 degrees C and evaporative heat loss was 132% of metabolic heat production. At high Ta there was only a slight increase in metabolic rate despite the employment of evaporative cooling mechanisms and an increase in Tb. Collectively our results suggest that M. condylurus is well suited to tolerate high Ta, and this may enable it to exploit thermally challenging roost sites and to colonise habitats and exploit food sources where less stressful roosts are limiting.     

A temperature-sensitive step in exocytosis.

We have examined the temperature sensitivity of exocytosis in digitonin-permeabilized chromaffin cells. The time course of secretion is markedly slowed by incubating the cells at 18 degrees C rather than 27 degrees C. We have previously shown that secretion has both ATP-dependent and ATP-independent components (Holz, R. W., Bittner, M. A., Peppers, S. C., Senter, R. A., and Eberhard, D. A. (1989) J. Biol. Chem. 264, 5412-5419). Reducing the temperature has no effect on ATP-independent secretion. However, cold (18 degrees C) greatly slows the ability of ATP to stimulate secretion. The ATP-requiring priming step itself is not affected by reducing the temperature since an effect of ATP can be seen after permeabilization at 18 degrees C if the cells are subsequently stimulated to secrete at 27 degrees C. When cells are permeabilized at 27 degrees C with ATP and then stimulated by Ca2+ in the absence of ATP, the secretion which was primed by ATP during the permeabilization step is inhibited 75% at 18 degrees C. Similar results are seen when ATP-dependent priming is enhanced by low concentrations of Ca2+. Thus, the temperature-sensitive step occurs after ATP and Ca2+ act to prime the cell. The temperature-sensitive step is likely to be overall rate-limiting step during the later phase of secretion, when the ATP-dependent priming process is limiting.     

The response of the high altitude C(4) grass Muhlenbergia montana (Nutt.) A.S. Hitchc. to long- and short-term chilling.

The acclimation of C(4) photosynthesis to low temperature was studied in the montane grass Muhlenbergia montana in order to evaluate inherent limitations in the C(4) photosynthetic pathway following chilling. Plants were grown in growth cabinets at 26 degrees C days, but at night temperatures of either 16 degrees C (the control treatment), 4 degrees C for at least 28 nights (the cold-acclimated treatment), or 1 night (the cold-stress treatment). Below a measurement temperature of 25 degrees C, little difference in the thermal response of the net CO(2) assimilation rate (A) was observed between the control and cold-acclimated treatment. By contrast, above 30 degrees C, A in the cold-acclimated treatment was 10% greater than in the control treatment. The temperature responses of Rubisco activity and net CO(2) assimilation rate were similar below 22 degrees C, indicating high metabolic control of Rubisco over the rate of photosynthesis at cool temperatures. Analysis of the response of A to intercellular CO(2) level further supported a major limiting role for Rubisco below 20 degrees C. As temperature declined, the CO(2) saturated plateau of A exhibited large reductions, while the initial slope of the CO(2) response was little affected. This type of response is consistent with a Rubisco limitation, rather than limitations in PEP carboxylase capacity. Stomatal limitations at low temperature were not apparent because photosynthesis was CO(2) saturated below 23 degrees C at air levels of CO(2). In contrast to the response of photosynthesis to temperature and CO(2) in plants acclimated for 4 weeks to low night temperature, plants exposed to 4 degrees C for one night showed substantial reduction in photosynthetic capacity at temperatures above 20 degrees C. Because these reductions were at both high and low CO(2), enzymes associated with the C(4) carbon cycle were implicated as the major mechanisms for the chilling inhibition. These results demonstrate that C(4) plants from climates with low temperature during the growing season can fully acclimate to cold stress given sufficient time. This acclimation appears to involve reversal of injury to the C(4) cycle following initial exposure to low temperature. By contrast, carbon gain at low temperatures generally appears to be constrained by the carboxylation capacity of Rubisco, regardless of acclimation time. The inability to overcome the Rubisco limitation at low temperature may be an inherent limitation restricting C(4) photosynthetic performance in cooler climates.     

A rapid screening method for the detection of viable spores in powder using bioluminescence.

A rapid diagnosis of a biological threat in a powder sample is important for fi rst responders who have to make decisions on-site. The present culture-based method does not provide timely results, which is a critical barrier for a quick response when a suspicious powder sample is found. The ATP bioluminescence method, combined with a heat shock, was investigated to determine the presence of spores in powder. The results show that only spore-containing powder samples provided a dramatic increase in the bioluminescence signal after the heat shock, which induces germination of the spores. Various conditions were tested to fi nd the most effective and rapid germination procedure. Elevated temperatures (37 degrees C and 50 degrees C) were more effective in germination than room temperature. At 50 degrees C, a double-strength germinant was more effective in germination than the regular strength. The 37 degrees C/15 min procedure induced the germination of spores most effectively, while a 50 degrees C/2 min procedure provided reasonably high signals, so it could make the entire procedure even faster (< 5 min). The detection limit of the bioluminescence method is < 100 spores.     

Heat shock-induced acquisition of thermotolerance at the levels of cell survival and translation in Xenopus A6 kidney epithelial cells.

In this study we have investigated the acquisition of thermotolerance in a Xenopus laevis kidney A6 epithelial cell line at both the level of cell survival and translation. In cell survival studies, A6 cells were incubated at temperatures ranging from 22 to 35 degrees degrees C for 2 h followed by a thermal challenge at 39 degrees degrees C for 2 h and a recovery period at 22 degrees C for 24 h. Optimal acquisition of thermotolerance occurred at 33 degrees degrees C. For example, exposure of A6 cells to 39 degrees degrees C for 2 h resulted in only 3.4% survival of the cells whereas prior exposure to 33 degrees C for 2 h enhanced the survival rate to 69%. This state of thermotolerance in A6 cells was detectable after 1 h at 33 degrees C and was maintained even after 18 h of incubation. Cycloheximide inhibited the acquisition of thermotolerance at 33 degrees C suggesting the requirement for ongoing protein synthesis. The optimal temperature for the acquisition of translational thermotolerance also occurred at 33 degrees C. Treatment of A6 cells at 39 degrees C for 2 h resulted in an inhibition of labeled amino acid incorporation into protein which recovered to approximately 14% of control after 19 h at 22 degrees C whereas cells treated at 33 degrees C for 2 h prior to the thermal challenge recovered to 58% of control levels. These translationally thermotolerant cells displayed relatively high levels of the heat shock proteins hsp30, hsp70, and hsp90 compared to pretreatment at 22, 28, 30, or 35 degrees C. These studies demonstrate that Xenopus A6 cells can acquire a state of thermotolerance and that it is correlated with the synthesis of heat shock proteins.     

Effect of sodium chloride and citric acid on growth and toxin production by A. caviae and A. sobria at moderate and low temperatures

The effect of sodium chloride and citric acid on hemolysin and caseinase production by Aeromonas caviae and Aeromonas sobria at 32 degrees C and 5 degrees C was investigated. At 32 degrees C, although both strains were tolerant to 3% NaCl in TSB, the production of caseinase was decreased in the presence of 1-3% NaCl, and the production of hemolysin was abolished by 2-3% NaCl. Citric acid (0.03%) was less effective than NaCl in reducing hemolysin and caseinase production by both strains at 32 degrees C. A combination of low temperature (5 degrees C) and citric acid treatment reduced hemolysin and caseinase production by both strains. A combination of low temperature (5 degrees C) and NaCl (3%) treatment was the most effective procedure in reducing growth and hemolysin and caseinase production by the tested strains.     

镰形臂尾轮虫和尾突臂尾轮虫的生活史特征比较

应用单个体培养方法,以浓度为3.0×10⁶ cells·ml^-1的斜生栅藻为食物,在18 ℃、23 ℃、28 ℃和33 ℃的温度梯度下比较了镰形臂尾轮虫和尾突尾轮虫的生活史特征.结果表明：18 ℃和23 ℃下,尾突臂尾轮虫的生殖期和平均寿命均显著长于镰形臂尾轮虫,产卵量也显著大于镰形臂尾轮虫;28 ℃下,2种轮虫的各主要发育阶段历时、平均寿命和产卵量均无显著差异;33 ℃下,镰形臂尾轮虫的生殖期和生殖后期历时以及平均寿命均显著长于尾突臂尾轮虫,产卵量显著大于尾突臂尾轮虫.18 ℃下,尾突臂尾轮虫的生命期望、净生殖率和种群内禀增长率均显著大于镰形臂尾轮虫;23 ℃和28 ℃下,尾突臂尾轮虫的生命期望显著长于镰形臂尾轮虫,而其他种群增长参数间均无显著差异;33 ℃下,镰形臂尾轮虫的世代时间、生命期望、净生殖率和种群内禀增长率均极显著大于尾突臂尾轮虫.2种轮虫的主要发育阶段历时、平均寿命、产卵量、世代时间、生命期望、净生殖率和种群内禀增长率对温度变化的反应也存在差异.研究结果表明,尾突臂尾轮虫更能适应较低的环境温度,而镰形臂尾轮虫则相反.     

Influence of postirradiation incubation temperature on recovery of radiation-injured Clostridium botulinum 62A spores.

The number of colonies formed by unirradiated Clostridium botulinum 62A spores was independent of temperature, in the range from 20 to 45 degrees C (in 5 degrees C increments); no colonies developed at 50 degrees C. Spores irradiated at 1.2 or 1.4 Mrads produced more macrocolonies at 40 degrees C than at higher or lower temperatures. Apparently, radiation-injured spores were capable of repair of 40 degrees C than at the other temperatures studied. More than 99% of the radiation (1.2 Mrads) survivors were injured and were unable to form macrocolonies in the presence of 5% NaCl. The germinated radiation-injured spores were also sensitive to dilution, resulting in the loss of viability of 77 to 79% of the radiation survivors. At 30 and 40 degrees C, the irradiated spores did not differ significantly in the extent of germination (greater than 99% at both 30 and 40 degrees C), emergence (64% at 30 degrees C and 67% at 40 degrees C), and the maximum number of emerged cells that started to elongate (69% at 30 degrees C and 79% at 40 degrees C). However, elongation was remarkably more extensive at 40 degrees C than at 30 degrees C. Many elongated cells lysed within 48 h at 30 degrees C, indicating an impaired repair mechanism. If the radiation-injured spores were incubated at 40 degrees C in the recovery (repair) medium for 8 to 10 h, they germinated, emerged, and elongated extensively and were capable of repair. If, after 8 to 10 h at 40 degrees C, these cultures were shifted to 30 degrees C, the recovery at 30 increased by more than eightfold, resulting in similar colony counts at 30 and 40 degrees C. Thus, repair appeared to be associated with outgrowth. Repair did not occur in the presence of chloramphenicol at 40 degrees C, whereas penicillin had no effect, suggesting that the repair involved protein synthesis but did not require multiplication.     

Effects of low culture temperature on the induction of hsp70 mRNA and the accumulation of hsp70 and hsp105 in mouse FM3A cells.

We have shown that heat shock does not induce the synthesis of hsp70 in FM3A cells maintained at a low culture temperature of 33 degrees C although it does so in cells maintained at 37 degrees C [T. Hatayama et al. (1991) Biochem. Int. 24, 467-474]. In this paper, we show that FM3A cells maintained at 37 degrees C produced hsp70 mRNA during continuous heating at 42 degrees C or during postincubation at either 37 or 33 degrees C after being heated at 45 degrees C for 15 min, whereas cells maintained at 33 degrees C did not produce hsp70 mRNA during continuous heating at 37, 39, 42, or 45 degrees C, or during postincubation after being heated at any temperature. Thus the lack of hsp70 synthesis in cells maintained at 33 degrees C seemed to be due to the absence of hsp70 mRNA induction. Also, hsp70 was accumulated in cells maintained at 37 degrees C during continuous heating at 42 degrees C and during postincubation at 37 degrees C after heat shock at 45 degrees C, but not during postincubation at 33 degrees C. The cellular level of the constitutive hsp73 as well as the mRNA level were both similar in cells maintained at 33 and 37 degrees C. On the other hand, the cellular level of the constitutive hsp105 in cells maintained at 33 degrees C was only half of that in cells maintained at 37 degrees C. These hsp105 levels increased significantly in both types of cells after continuous heating at 39 degrees C. These findings indicate that the culture temperature affects not only the induction of hsp70 mRNA but also the accumulation of hsp70 and hsp105 in the cells.     

Effect of protein disulfide isomerase on the regeneration of bovine ribonuclease A with dithiothreitol.

The role of protein disulfide isomerase (PDI) in the regeneration of ribonuclease A with dithiothreitol (DTT) was investigated at three different temperatures. The rates of formation of the native protein were markedly increased in the presence of PDI, 9-fold at 15 degrees C, 6-fold at 25 degrees C and 62-fold at 37 degrees C, respectively. In the presence of PDI, major changes were found in the distribution of intermediates in the three-disulfide region at 25 and 15 degrees C and also in the one-disulfide region at 15 degrees C, with the fast accumulation of the two native-like species des-[65-72] and des-[40-95]. The present results indicate that PDI does not alter the two major parallel pathways involving des-[65-72] and des-[40-95] in the regeneration of ribonuclease A with DTT.     

A transient-kinetic study of the nitrogenase of Klebsiella pneumoniae by stopped-flow calorimetry. Comparison with the myosin ATPase.

The pre-steady-state kinetics of MgATP hydrolysis by nitrogenase from Klebsiella pneumoniae were studied by stopped-flow calorimetry at 6 degrees C and at pH 7.0. An endothermic reaction (delta Hobs. = +36 kJ.mol of ATP-1; kobs. = 9.4 s-1) in which 0.5 proton.mol of ATP-1 was released, has been assigned to the on-enzyme cleavage of MgATP to yield bound MgADP + Pi. The assignment is based on the similarity of these parameters to those of the corresponding reaction that occurs with rabbit muscle myosin subfragment-1 (delta Hobs. = +32 kJ.mol of ATP-1; kobs. = 7.1 s-1; 0.2 proton released.mol of ATP-1) [Millar, Howarth & Gutfreund (1987) Biochem. J. 248, 683-690]. MgATP-dependent electron transfer from the nitrogenase Fe-protein to the MoFe-protein was monitored by stopped-flow spectrophotometry at 430 nm and occurred with kobs. value of 3.0 s-1 at 6 degrees C. Thus, under these conditions, hydrolysis of MgATP precedes electron transfer within the protein complex. Evidence is presented that suggests that MgATP cleavage and subsequent electron transfer are reversible at 6 degrees C with an overall equilibrium constant close to unity, but that, at 23 degrees C, the reactions are essentially irreversible, with an overall equilibrium constant greater than or equal to 10.     

A comparative study of the thermal stability of oligodeoxyribonucleotides containing 5-substituted 2'-deoxyuridines.

Two series of modified oligonucleotides based on the self-complementary dodecamer d(CGCTAATTAGCG) were synthesized. The first contained the -C identical withCCH2R linker at C5 of deoxyuridine at position 4 (T*) of d(CGCT*AATTAGCG) and the second contained the -SR linker. The goal of the study was to evaluate and compare these two types of side chains for suitability as tethers for linking reporter groups to oligonucleotides. Our primary concern was how these tethers would effect duplex stability. The modified nucleosides were synthesized by palladium-mediated coupling reactions between the substituted alkyne and 5'-(4, 4'-dimethoxytrityl)-5-iodo-2'-deoxyuridine and between a disulfide and 5-chloromercurio-2'-deoxyuridine. The C5 deoxyuridine side chains evaluated included C identical with CCH3, C identical with CCH2NHC(O)CH3, C identical with CCH2N(CH3)2, C identical with CCH2N-HC(O)C5H4N, C identical with CCH2NHC(O)C10H15, SCH3, SC6H5 and SCH2CH2NHC(O)CH3. The nucleosides containing these substituents were incorporated into oligo-deoxyribonucleotides by standard phosphoramidite methodology. Melting studies demonstrated that the sequence containing the C identical with CCH3side chain had the highest T m value (59.1 degrees C) in comparison with the control sequence (T m = 55.2 degrees C) and that any additional substituent on C3 of the propynyl group lowered the T m value relative to propynyl. Nevertheless, even the most destabilizing substituent, adamantylcarbamoyl, yielded an oligodeoxyribonucleotide that dissociated with a T m of 54 degrees C, which is only 1.2 degrees C less than the control sequence. In contrast, the thioether substituents led to lower T m values, ranging from as low as 45.1 degrees C for SPh up to 52.2 degrees C for SMe. Replacing the methyl of the SMe substituent with a CH2CH2NHC(O)CH3 tether led to no further reduction in melting temperature. The T m value of the CH2CH2NHC(O)CH3-containing oligonucleotide was less than the natural sequence by 1.6 degrees C/substituent. This is sufficiently small that it is anticipated that the C5 thioether linkage may be as useful as the acetylenic linkage for tethering reporter groups to oligonucleotides. More importantly, the thioether linkage provides a means to position functional groups to interact specifically with opposing complementary (target) sequences.     

Hydroxymethylglutaryl-coenzyme A reductase. Purification and properties of the enzyme from Fusarium oxysporum.

The hydroxymethylglutaryl-coenzyme A reductase (mevalonate:NADP+ oxidoreductase, EC 1.1.1.34) system in Fusarium oxysporum, a soil inhabiting plant pathogen, has been examined. Two forms of the enzyme catalyzing the conversion of hydroxymethylglutaryl-coenzyme A were obtained in the supernatant after precipitation at 75% (NH4)2SO4 saturation of the soluble culture extract which was previously separated from cell wall, mitochondria and microsomes. The two forms of the enzyme were separated electrophoretically. A third form, contained in the precipitate obtained at 35--75% (NH4)2SO4 saturation of the same extract, was further purified by Sephadex G-50 column chromatography. This purified form moved as a single band in sodium dodecyl sulphate electrophoresis and in immunological tests and has a molecular weight of 11 000. The apparent Michaelis constant for the substrate hydroxymethylglutaryl-coenzyme A is 21 micron at 2 micron NADP. NADPH is a more efficient reductant on a molar basis than NADH for the deacylation of the hydroxymethylglutaryl-coenzyme A substrate. Optimum activity of the enzyme was obtained at pH 7.4 and 37 degrees C. The enzyme demonstrated no cold sensitivity but rather was more stable at 4 degrees C than at 25 degrees C. The protection with dithiothreitol, though minimal compared to other systems, was more effective at the higher temperature.     

Membrane binding induces lipid-specific changes in the denaturation profile of bovine prothrombin. A scanning calorimetry study.

Prothrombin denaturation was examined in the presence of Na2EDTA, 5mM CaCl2, and CaCl2 plus membranes containing 1-palmitoyl-2-oleoyl-3-sn-phosphatidylcholine (POPC) in combination with either bovine brain phosphatidylserine (PS) or 1,2-dioleoyl-phosphatidylglycerol (DOPG). Heating denaturation of prothrombin produced thermograms showing two peaks, a minor one at approximately 59 degrees C previously reported to correspond to denaturation of the fragment 1 region (Ploplis, V. A., D. K. Strickland, and F. J. Castellino 1981. Biochemistry. 20:15-21), and a main one at approximately 57-58 degrees C, reportedly due to denaturation of the rest of the molecule (prethrombin 1). The main peak was insensitive to the presence of 5mM Ca2+ whereas the minor peak was shifted to higher temperature (Tm approximately 65 degrees C) by Ca2+. Sufficient concentrations of POPC/bovPS (75/25) large unilamellar vesicles to guarantee binding of 95% of prothrombin resulted in an enthalpy loss in the main endotherm and a comparable enthalpy gain in the minor endotherm accompanying an upward shift in peak temperature (Tm approximately 73 degrees C). Peak deconvolution analysis on the prothrombin denaturation profile and comparison with isolated prothrombin fragment 1 denaturation endotherms suggested that the change caused by POPC/PS vesicles reflected a shift of a portion of the enthalpy of the prethrombin 1 domain to higher temperature (Tm approximately 77 degrees C). The enthalpy associated with this high-temperature endotherm increased in proportion to the surface concentration of PS. By contrast, POPC/DOPG (50/50) membranes shifted the prethrombin 1 peak by 4 degrees C to a lower temperature and the fragment 1 peak by 5 degrees C to a higher temperature. The data lead to a hypothesis that the fragment 1 and prethrombin 1 domains of prothrombin do not denature quite independently and that binding of prothrombin to acidic-lipid membranes disrupts the interaction between these domains. It is further hypothesized that PS containing membranes exert the additional specific effect of decoupling the denaturation of two subdomains of the prethrombin 1 domain of prothrombin.     

A novel ribonuclease from fresh fruiting bodies of the portabella mushroom Agaricus bisporus.

A 14 kDa ribonuclease with a novel N-terminal sequence was isolated from fresh fruiting bodies of the portabella mushroom. It was adsorbed on DEAE-cellulose and carboxymethyl-cellulose, and demonstrated the highest ribonucleolytic potency toward poly (A), 60% as much activity toward poly (C), 40% as much activity toward poly (U), and the least activity (7% as much) toward poly (G). It exhibited a pH optimum at pH 4.5 and a temperature optimum at 60 degrees C. Its activity at 100 degrees C was higher than that at 20 degrees C.     

A comparison of physiological changes in carp, Cyprinus carpio, induced by several pollutants at sublethal concentration--II. The dependency on the temperature

Carp were exposed to 10 different pollutants with sublethal concentrations at 12, 17 or 22 degrees C. The effects on the serum cortisol and glucose levels, the amount of liver and muscle glycogen and the concentration of protein and cholesterol in the serum were examined. The level of serum cortisol and glucose increased, the amount of liver and muscle glycogen decreased and the protein and cholesterol concentration were reduced after exposure to the pollutants. The cortisol and glucose response was similar at all temperatures, slightly reduced at 17 degrees C. The glycogen reaction was strongest at 17 degrees C and the protein and cholesterol response was low at 12 degrees C and increased with the temperature. The explanation of the different temperature dependency is difficult. Carp are stressed and have low metabolic rate at 12 degrees C. They have good conditions at 17 degrees C and have a higher energy demand at 22 degrees C. These simple differences cannot exactly explain the different dependency.