Thermodynamic and kinetic study of thermal denaturation of Kunitz soybean trypsin inhibitor by differential scanning microcalorimetry

The thermal denaturation of soybean trypsin inhibitor (Kunitz inhibitor) has been studied in pH-region from 2.0 to 11.0 by differential scanning microcalorimetry. The thermodynamic characteristics have been determined. It has been established that the denaturation transition of protein may be described by a two-state model. It has been shown, that two side hydrogen bonds between carboxylate-ion and tyrosyl and carboxylate-ion and lysyl take part in the stabilization of the inhibitor's native structure. The activation of denaturation is accompanied by cleavage of one side hydrogen bond.     

血管内皮舒张因子在氧自由基所致慢性缺氧大鼠肺内动脉收缩中的作用

以黄嘌岭(X)-黄嘌呤氧化酶(XO)系统产生氧自由基,应用微量生物测定法观察慢性缺氧(5000m,10d)对大鼠氧自由基所致肺内动脉收缩的影响及内皮舒张因子(EDRF)在其中的作用。慢性缺氧大鼠有内皮的肺内动脉环对氧自由基的收缩反应较正常环境中的对照动物明显增强,加入EDRF灭活剂还原型血红蛋白(RHb)后更加显著;而加入超氧化物歧化酶(铜锌SOD)后则减弱,甚至消除。反之,不论加入RHb或SOD对氧自由基所致去内皮肺内动脉环的收缩反应均无明显影响。上述结果表明慢性缺氧引起肺内动脉收缩增强与EDRF有密切关系:慢性缺氧可能使EDRF的作用减弱,肺内动脉对氧自由基的反应性增强。表示EDRF及其与氧自由基的关系在慢性缺氧性肺动脉高压的形成中可能具有十分重要的意义。     

Thiol-induced oligomerization of α-lactalbumin at high pressure

Denaturation and aggregation of-lactalbumin at high pressure (up to 10 kbar, 1000 MPa) were studied by means of circular dichroism, gel-permeation chromatography, sodium dodecyl sulfate and gel electrophoresis. It was found that the unfolding of-lactalbumin at high pressure is reversible even in basic pH and at a protein concentration as large as 10%. In these conditions only a negligible fraction of the protein is denatured irreversibly and aggregates. The rate of aggregation of-lactalbumin at high pressure increases significantly in the presence of low-molecular reducing agents such as cysteine, 2-mercaptoethanol, and dithiothreitol. Maximal yield of-lactalbumin oligomerization (over 90%) was achieved in the presence of cysteine at the molar cysteine/protein ratioq=2 and atpH 8.5. Apparent molecular weight of the obtained oligomers was over 500 kDa. It was shown that the size distribution of oligomers can be modulated by varyingpH and reducing agent. The size distribution shifts in the direction of very large, poorly soluble particles whenpH decreases. Maximal content of the insoluble fraction (about 30%) can be reached at pH 5.5 when cysteine (q=2) is used as reducing agent. The oligomers of-lactalbumin are stabilized mainly by nonnative interchain disulfide bridges. Circular dichroism measurements point to an additional mechanism of cohesion of polypeptide chains in the oligomers, which is formation of intermolecular-sheets.     

Use of polysaccharides to remove lipids from the protein globulin fraction of baker's yeast

The precipitation of lipid-protein complexes from the baker's yeast protein globulin fraction by polysaccharides (gum arabic and arabinogalactan) was investigated. Lipid-protein complexes were precipitated more readily with the polysaccharides under study than with other globulin fractions components. A method for the removal of lipids from the globulin fraction of baker's yeast by precipitation of the lipid-protein complexes with polysaccharides is suggested.     

Determination of nystatin component composition using HPLC and TLC with densitometry]

The component composition of nystatin produced by an improved strain of Streptomyces noursei was determined by HPLC on Milichrom chromatograph (USSR). It was shown that the antibiotic consisted of nystatins A1, A2, A3 and B and admixture substances. The data appeared to be in good agreement with the results of the complex TLC investigation, by using densitometry. The component composition of the samples was evidenced by SIEAP mass spectrometry. Physiochemical and biological characteristics of separate components are presented.     

Early Diagnosis of Neurodegenerative Diseases – The Long Awaited Holy Grail and Bottleneck of Modern Brain Research – 19th HUPO BPP Workshop

The HUPO Brain Proteome Project (HUPO BPP) held its 19th workshop in Dortmund, Germany, from May 22 to 24, 2013. The focus of the spring workshop was on strategies and developments concerning early diagnosis of neurodegenerative diseases     

Calorimetric study of the order-disorder conformational transition in succinoglycan

Thermally induced order-disorder conformational transition in succinoglycan was studied using the method of high-sensitivity differential scanning microcalorimetry within the range of polysaccharide concentrations from 0.1 to 3.5 mg mL⁻¹ at NaCl concentrations 0, 0.01, and 0.1M. The positions and shapes of the excess heat capacity curves depended substantially on both the NaCl and polysaccharide concentrations. At low polysaccharide concentrations in salt-free solution the experimental curves were closely approximated by the two-state model suggesting the transition mechanism to be of the single helix-coil type. With increasing polysaccharide and/or NaCl concentration, the experimental curves changed significantly in symmetry, which indicated a changing transition mechanism. At high polysaccharide concentrations or in the presence of the salt, the order-disorder transition of succinoglycan was shown to include two stages: the cooperative dissociation of the helix dimer and subsequent two-state melting of the helix monomer. The dependence of thermodynamic parameters for the dissociation and melting of helix structures in succinoglycan on NaCl and polysaccharide concentrations was obtained by fitting the experimental excess heat capacity curves. The cooperativity parameter σ for the single helix-coil transition as well as the average length of the helix segment of succinoglycan were calculated. Some features of succinoglycan ordering in solution are discussed. © 1996 John Wiley & Sons, Inc.     

Use of subcritical fluid chromatography for the separation of enantiomers using packed cellulose based stationary phase

Investigations into the parameters affecting the enantiomeric separation of an intermediate in the synthesis of a drug targeted for cardiac arrhythmia is presented. The separation was achieved under subcritical fluid chromatography using CO₂ modified with methanol and co-modifiers isopropyl amine and methylene chloride. The stationary phase was a cellulose based stationary phase manufactured under the trade name Chiracel OB. The results showed the addition of methylene chloride had a noticeable effect on resolution while the separation factor, α, remained constant. The co-modifier, isopropyl amine, did not considerably influence the resolution, or separation factor. Temperature studies were performed in order to establish the thermodynamic parameters of the interaction between the enantiomers and the stationary phase. Plots of In k′ vs. 1/T proved to be curved, while plots of In α vs. 1/T were linear. © 1996 Wiley-Liss, Inc.