红细胞膜上结合水的萤光与红外研究

水是生物膜上重要的结构与功能分子。本实验以20℃红细胞膜对水的等温吸附曲线为定量根据,研究了不同水化水平的水化效应。DPH的萤光偏振测量证明:脱水将破坏脂双层的有序结构,大约20%的水化度是维持膜结构所必需的;在膜结构完整的条件下结合水的增加会使膜流动性增大,水分子对双层类脂分子的“松弛效应”和水分子的运动对于膜脂分子的活动性可能有重要贡献。ANS的萤光发射谱说明水化效应增加了膜表面的极性,也提示在高水化水平膜上存在着一部份活动性较大的水分子。红外差示谱表明膜上存在着两类结合水——紧密结合水与松弛结合水,也说明脱水将造成磷脂与胆固醇的相分离现象。     

石英粉尘对人红细胞膜的损伤及柠檬酸铝的抗损伤作用研究——V、对膜蛋白构象的影响

本研究用圆二色技术测定石英粉尘对人红细胞膜蛋白构象的影响以及柠檬酸铝拮抗石英影响的效应.结果显示,石英引起膜蛋白构象改变较为迅速;对膜蛋白构象的影响(α-螺旋量减少)存在着剂量效应关系.柠檬酸铝可以有效地减轻石英对红细胞膜蛋白构象的影响.     

石英粉尘对人红细胞膜的损伤及柠檬酸铝的抗损伤作用研究——V、对膜蛋白构象的影响

本研究用圆二色技术测定石英粉尘对人红细胞膜蛋白构象的影响以及柠檬酸铝拮抗石英影响的效应.结果显示,石英引起膜蛋白构象改变较为迅速;对膜蛋白构象的影响(α-螺旋量减少)存在着剂量效应关系.柠檬酸铝可以有效地减轻石英对红细胞膜蛋白构象的影响.     

脱水在聚乙二醇诱导膜融合中的作用

随着各种诱导膜融合的因子相继发现,人们建立了各种膜融合的模型.我们通过对聚乙二醇PEG诱导脂质体融合的分析,认为膜融合的关键在于脱去膜表面的结合水,而其它作用诸如膜脂缺陷.膜脂分相以及脂多型性等尽管是不同膜体系中直接观察到的膜融合形式,都是膜脱去结合水带来的必然结果.膜表面结合水的排除是前因,本文着重讨论脱水及脱水后膜脂结构的一系列变化.     

二氧化硅及柠檬酸铝对巨噬细胞膜K~+通透性影响的研究

通常认为矽肺发病的重要环节之一是肺泡巨噬细胞吞噬二氧化硅(SiO_2)粉尘颗粒后的崩解死亡。因此,凡能保护巨噬细胞的药物就可能具有防治矽肺的作用。柠檬酸铝(柠铝)对实验性动物矽肺以及临床矽肺病人的疗效可能正是与其体外保护巨噬细胞的作用有关。最近体外研究证明,SiO_     

温石棉对人红细胞膜脂质及蛋白质影响的ESR研究

用５ＮＳ、１６ＮＳ及ＭＳＬ标记人红细胞膜,观察了茫崖及涞温源石棉对膜脂质及蛋白质的影响。结果表明,两种温石棉均可增加膜表面层及深层脂质刚性,即致膜表、深层脂质流动性降低;同时可改变膜蛋白构象。两种温石棉时膜脂质及蛋白质的影响强度与其剂量有关。经柠檬酸铝处理后,温石棉的上述作用明显减弱。     

补体与红细胞膜的结合作用

被激活的补体3(C_3),能与红细胞膜结合;C_(3b)与红细胞膜形成的复合物,又激活一系列补体(C_5-C_9),使细胞溶解。有些溶血性疾病与此种补体溶血有关。我们实验曾证明,阵发性睡眠性血红蛋白尿症(PNH)红细胞对补体溶血敏感。为了探讨PNH补体溶血反应的机理,我们曾用对唾液酸专一结合的鲎血凝集素处理PNH红细胞,表明它能降低补体溶血,并证明它与红细胞膜血型糖蛋白结合。看来鲎血凝集素可能影响红细胞膜与C_(3b)的结     

山莨菪碱对人红细胞膜蛋白及膜脂运动的影响

本文采用自旋标记顺磁共振波谱技术,研究了山茛菪碱对人红细胞膜蛋白和膜脂运动的影响.结果表明:用马来酰亚胺标记的人红细胞膜,加入山茛菪碱后,其顺磁共振波谱中强、弱固定化作用谱的峰值比增大,膜蛋白的运动受到限制.山茛菪碱对红细胞膜脂的作用部位主要在极性头部,并影响膜脂的流动性.本文还对山茛菪碱与红细胞膜作用的可能机制进行了讨论.     

吗啡对大鼠红细胞膜生物物理特性的影响

为研究吗啡对生物体血液系统的影响,以大鼠为研究对象,分别培养了三周成瘾的吗啡依赖模型、体外吗啡作用和吗啡急性注射的模型,研究了吗啡对大鼠红细胞膜的生物物理特性的改变。作者采用新型激光衍射法测量了上述血样的弹性模量Ｅ和膜微粘度（μｍ）,同时通过ＤＰＨ标记的荧光偏振法测定了这些红细胞膜的流动性,还采用傅立叶变换的红外技术（ＦＴ－ＩＲ）测量了红细胞膜蛋白构象的动态变化,并对上述红细胞膜结构改变所引起的微观流变特性的变化进行了初步探讨。结果表明,大鼠对吗啡形成依赖会导致其红细胞膜流动性的下降,弹性剪切模量的升高和膜蛋白质二级结构的变化。而吗啡的体外作用和体内急性作用对大鼠红细胞膜特性的影响与吗啡依赖的作用相类似。由此我们推测吗啡对大鼠红细胞膜特性的影响可能不需通过受体,而是直接发生作用的。     

二氧化硅对细胞膜的损伤和柠檬酸铝抗损伤作用机理的拉曼光谱研究

 本研究应用激光拉曼散射光谱技术探讨SiO_2了诱发膜损伤效应以及柠檬酸铝抗损伤效应的机理。提出SiO_2的作用机理主要是与膜脂极性头部基团-N~+(CH_3)_3的相互作用及其对脂双层有序结构的破坏;而柠檬酸铝的抗损伤作用机理主要是通过铝与SiO_2颗粒表面的结合,从而阻断SiO_2与膜的直接相互作用,在一定程度上维持了膜的结构和功能。     

A Study of the Stationary Volumetric Elastic Modulus during Dehydration and Rehydration of Stems of Pea Seedlings

The relationship between cortical-cell turgor pressure (P) and tissue water mass (W) was determined for stem segments of pea (Pisum sativum L.) seedlings subjected to hydration and dehydration. This allowed a test for elastic hysteresis in the cortical cells. The P-W curves for dehydration and hydration were not coincident. In some experiments, the P-W curves exhibited a "roll-off" at high P, similar to the "plateau effect" sometimes observed in pressure-chamber studies. When hydration was followed by a 4-h dehydration, the tissue water mass (W0) at minimum turgor was reduced. This might reflect a reduction in apoplastic water mass and/or a contraction of the symplast during dehydration. Neglecting the decrease in W0 leads to underestimates of the stationary volumetric elastic modulus ([epsilon]stat). The result of an analysis that assumes W0 was constant during hydration suggests that there was no significant difference in [epsilon]stat between dehydration and hydration and, hence, no significant elastic hysteresis. However, a 16-h dehydration increased [epsilon]stat; this might be a response to water stress.     

脱水方法对棕榈种子萌发及膜脂过氧化的影响

以棕榈种子为材料,比较了硅胶脱水和自然脱水方法下种子萌发特征和膜脂过氧化程度。结果表明:棕榈种子的初始含水量为33.1%,萌发率为83.3%;当硅胶脱水至含水量21.2%时萌发率为80.0%,而自然脱水至23.2%时萌发率仅为56.7%;当含水量降至10%左右时,硅胶脱水萌发率为27.7%,而自然脱水的萌发率为26.7%。在脱水过程中,2种脱水处理种子的浸出液电导率和丙二醛(MDA)含量都呈升高趋势,但自然脱水种子浸出液电导率升高的速率较硅胶脱水快,而MDA含量在硅胶脱水下增加较大。硅胶脱水处理种胚中脯氨酸含量及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、过氧化物酶(POD)和抗坏血酸过氧化物酶(APX)活性均较自然脱水高,但2种脱水处理种子整体均呈先增加再下降的趋势。研究发现,棕榈种子为中间型种子,其在脱水初期对自然脱水较敏感,而脱水后期脱水速率对其生活力影响较小;棕榈种子对硅胶脱水的脱水敏感较自然性脱水要低,硅胶脱水有利于改善棕榈种子的贮藏寿命。     

Dehydration of cytosine monohydrate at physiological temperatures.

Neutron diffraction, thermogravimetric, and mass spectrographic measurements have been used to show that cytosine monohydrate loses its water of hydration at physiological temperatures (approximately equal to 37 degrees C) and converts to cytosine. The "activation energy" for the dehydration process has been determined from isothermal weight curves and is 27.1 +/- 0.6 kcal . mol-1. It is suggested that pyrimidine dehydration may be involved in structural changes in DNA.     

Age-dependent magnetosensitivity of heart muscle hydration

The reason for hyper magnetosensitivity of young animals compared to older ones remains unclear. It has been suggested that age-induced tissue dehydration (decreased water content) could be a basis for the aging-related decrease in the organism's magnetosensitivity. To test this hypothesis, the effect of a 0.2 T static magnetic field (SMF) exposure on heart muscle hydration in three age groups of rats (young, adult, and older) was studied, with and without ouabain poisoning. The SMF exposure resulted in heart muscle dehydration of young (21%) and adult (6.2%) rats but had no effect on older animals. In young animals without ouabin poisoning, SMF exposure caused dehydration of the heart muscle while in the ouabain-poisoned animals it led to hydration (29.6%). These hydration effects were more pronounced in young animals than in adult and older animals. The increased hydration (5.7%) of heart muscles in older animals was evoked by providing distilled water for seven days, which elevated (by 12%) the SMF-induced heart muscle hydration effect. These results suggest that the hyper magnetosensitivity of the young heart muscle and the lower sensitivity of older animals are due to initial high (83.5%) and low (75.3%) tissue hydration levels, respectively. Therefore, the age-induced decrease in the magnetosensitivity of heart muscle is likely to be a result of Na(+)/K(+) pump dysfunction.     

Effect of dioxane on the structure and hydration-dehydration of alpha-chymotrypsin as measured by FTIR spectroscopy

A new experimental approach based on FTIR spectroscopic measurements was proposed to study simultaneously the adsorption/desorption of water and organic solvent on solid enzyme and corresponding changes in the enzyme secondary structure in the water activity range from 0 to 1.0 at 25 degrees C. The effect of dioxane on the hydration/dehydration and structure of bovine pancreatic alpha-chymotrypsin (CT) was characterized by means of this approach. Dioxane sorption exhibits pronounced hysteresis. No sorbed dioxane was observed at low water activities (a(w)<0.5) during hydration. At a(w) about 0.5, a sharp increase in the amount of sorbed dioxane was observed. Dioxane sorption isotherm obtained during dehydration resembles a smooth curve. In this case, CT binds about 150 mol dioxane/mol enzyme at the lowest water activities. Three different effects of dioxane on the water binding by the initially dried CT were observed. At a(w)<0.5, water adsorption is similar in the presence and absence of dioxane. It was concluded that the presence of dioxane has little effect on the interaction between enzyme and tightly bound water at low a(w). At a(w)>0.5, dioxane increases the amount of water bound by CT during hydration. This behavior was interpreted as a dioxane-assisted effect on water binding. Upon dehydration at low water activities, dioxane decreases the water content at a given a(w). This behavior suggests that the suppression in the uptake of water during dehydration may be due to a competition for water-binding sites on chymotrypsin by dioxane. Changes in the secondary structure of CT were determined from infrared spectra by analyzing the structure of amide I band. Dioxane induced a strong band at 1628 cm(-1) that was assigned to the intermolecular beta-sheet aggregation. Changes in the intensity of the 1628 cm(-1) band agree well with changes in the dioxane sorption by CT. An explanation of the dioxane effect on the CT hydration and structure was provided on the basis of hypothesis on water-assisted disruption of polar contacts in the solid enzyme. The reported results demonstrate that the hydration and structure of alpha-chymotrypsin depend markedly on how enzyme has been hydrated - whether in the presence or in the absence of organic solvent. A qualitative model was proposed to classify the effect of hydration history on the enzyme activity-a(w) profiles.     

Kinetics and product analysis of the reaction catalysed by recombinant homoaconitase from Thermus thermophilus

HACN (homoaconitase) is a member of a family of [4Fe-4S] cluster-dependent enzymes that catalyse hydration/dehydration reactions. The best characterized example of this family is the ubiquitous ACN (aconitase), which catalyses the dehydration of citrate to cis-aconitate, and the subsequent hydration of cis-aconitate to isocitrate. HACN is an enzyme from the alpha-aminoadipate pathway of lysine biosynthesis, and has been identified in higher fungi and several archaea and one thermophilic species of bacteria, Thermus thermophilus. HACN catalyses the hydration of cis-homoaconitate to (2R,3S)-homoisocitrate, but the HACN-catalysed dehydration of (R)-homocitrate to cis-homoaconitate has not been observed in vitro. We have synthesized the substrates and putative substrates for this enzyme, and in the present study report the first steady-state kinetic data for recombinant HACN from T. thermophilus using a (2R,3S)-homoisocitrate dehydrogenase-coupled assay. We have also examined the products of the reaction using HPLC. We do not observe HACN-catalysed 'homocitrate dehydratase' activity; however, we have observed that ACN can catalyse the dehydration of (R)-homocitrate to cis-homoaconitate, but HACN is required for subsequent conversion of cis-homoaconitate into homoisocitrate. This suggests that the in vivo process for conversion of homocitrate into homoisocitrate requires two enzymes, in simile with the propionate utilization pathway from Escherichia coli. Surprisingly, HACN does not show any activity when cis-aconitate is substituted for the substrate, even though other enzymes from the alpha-aminoadipate pathway can accept analogous tricarboxylic acid-cycle substrates. The enzyme shows no apparent feedback inhibition by L-lysine.     

Cyclic AMP-dependent signaling system is a primary metabolic target for non-thermal effect of microwaves on heart muscle hydration

Previously, we have suggested that cell hydration is a universal and extra-sensitive sensor for the structural changes of cell aqua medium caused by the impact of weak chemical and physical factors. The aim of present work is to elucidate the nature of the metabolic messenger through which physiological solution (PS) treated by non-thermal (NT) microwaves (MW) could modulate heart muscle hydration of rats. For this purpose, the effects of NT MW–treated PS on heart muscle hydration, [³H]-ouabain binding with cell membrane, ⁴⁵Ca²⁺ uptake and intracellular cyclic nucleotides contents in vivo and in vitro experiments were studied. It is shown that intraperitoneal injections of both Sham-treated PS and NT MW–treated PS elevate heart muscle hydration. However, the effect of NT MW–treated PS on muscle hydration is more pronounced than the effect of Sham-treated PS. In vitro experiments NT MW–treated PS has dehydration effect on muscle, which is not changed by decreasing Na⁺ gradients on membrane. Intraperitoneal injection of Sham- and NT MW–treated PS containing ⁴⁵Ca²⁺ have similar dehydration effect on muscle, while NT MW–treated PS has activation effect on Na⁺/Ca²⁺ exchange in reverse mode. The intraperitoneal injection of NT MW–treated PS depresses [³H]-ouabain binding with its high-affinity membrane receptors, elevates intracellular cAMP and decreases cGMP contents. Based on the obtained data, it is suggested that cAMP-dependent signaling system serves as a primary metabolic target for NT MW effect on heart muscle hydration.     

Transition processes in stratum corneum model lipid membranes with a mixture of free fatty acids

The hydration of model membranes based on ceramide 6 with a mixture of free fatty acids most commonly encountered in the native lipid matrix of stratum corneum, the outermost layer of the mammalian skin, has been studied by neutron diffraction. Membrane hydration with water vapor at a temperature of 25°C is characterized by a small increase in the repeat distance Δd ₀ = 1.0 Å, which is comparable with membrane swelling in the presence of excess water. The kinetics of changes in the repeat distance, connected with an increase of the water layer between bilayers during hydration, and water exchange during the processes of hydration and H-D isotopic substitution, consists of a fast initial and a subsequent slow stage and is well described by exponentials with two characteristic times lying in the range from a few tens of minutes to several hundreds of minutes. During hydration at a temperature of 57°C, the repeat distance increases by Δd ₀ = 1.6 Å, after which the membrane irreversibly separates into two phases. One of the phases is formed mainly by long-chain free fatty acids and is characterized by a large decrease in the repeat distance Δd _ph = 8.3 Å on dehydration. The investigation of the structure of model membranes in the temperature range 20–72°C indicated that the system with 20% (w) of cholesterol in the range of 63–67°C undergoes a structural phase transition caused by the melting of hydrocarbon chains of lipids. In the system with a smaller content of cholesterol, no phase transition was observed up to a temperature of 72°C.