Detection of the associated state of membrane proteins by polyacrylamide gradient gel electrophoresis with non-denaturing detergents Application to band 3 protein from erythrocyte membranes

Polyacrylamide gradient gel electrophoresis was carried out in micellar solutions of various detergents which differ in degree of potency to denature proteins. From the application of this method to band 3 protein from erythrocyte membranes, it was suggested that the procedure was useful in studying the molecular state of membrane proteins.The electrophoretic behaviors of human and bovine band 3 protein did not show any species specificity in either a denature state and a state resembling the native state. As well as in nonionic detergent solutions, the dimeric and tetrameric structures of bovine band 3 protein were preserved in sodium deoxycholate solution, in which protein complexes maintained in nonionic detergent solutions are frequently dissociated. Even in dodecyltrimethylammonium bromide solution, which is a denaturant for water-soluble proteins, part of the band 3 protein was still present as the oligomer. The results suggest that the oligomeric form of band 3 protein is the stable structure and that the dimer and tetramer possibly coexist in membranes.     

Schistosoma mansoni: Electrophoretic characterization of strains selected for different levels of infectivity to snails

Individual adult Schistosoma mansoni from strains selected for high or low infectivity to specific strains of the snail intermediate host, Biomphalaria glabrata, were subjected to enzyme electrophoresis on starch gels. Fourteen enzyme systems were analyzed in an attempt to find electrophoretic markers associated with genes for infectivity to snails. The S. mansoni strains were selected from different isolates from Puerto Rico in several strains of B. glabrata. Of an estimated 18 loci, 3 were polymorphic and the remainder monomorphic. For 1 of the 3 polymorphic enzyme loci, lactate dehydrogenase (Ldh, EC 1.1.1.27), phenotype frequencies were correlated with infectivity to snails. In schistosome strains of low infectivity, frequencies of the Ldh-N phenotype ranged between 0.56 and 0.69, while in strains of high infectivity, Ldh-N frequencies were typically 0.91 to 1.00. Whether the correlation is accidental or due to some form of association, such as chromosomal linkage, between the locus responsible for variation in lactate dehydrogenase and a gene for infectivity to snails remains to be determined.     

Separation of algal mixtures and bacterial mixtures with flow-microfluorometer using chlorophyll and ethidium bromide fluorescence

The applicability of flow-microfluorometer to separate microbial cells was demonstrated with algal and bacterial cells. Algal mixtures were sorted according to the natural chlorophyll fluorescence and the bacterial mixtures were sorted according to the fluorescence of ethidium bromide-stained nucleic acid.Abbreviation FMF Flow-microfluorometer     

Surface potential and the interaction of weakly acidic uncouplers of oxidative phosphorylation with liposomes and mitochondria

The pH dependence of the binding of weakly acidic uncouplers of oxidative phosphorylation to rat-liver mitochondria and liposomes is mainly determined by the pK_a of the uncoupler molecule.

The absorption and fluorescence excitation spectra of the anionic form of weakly acidic uncouplers of oxidative phosphorylation are red-shifted upon interaction with liposomal or mitochondrial membranes. The affinity for the liposomes, as deduced from the red shift, is independent of the degree of saturation of the fatty acid chains of different lecithins. The intensity of the spectra at one pH value is strongly dependent upon the surface charge of the liposomes. With positively charged liposomes the results obtained can be almost quantitatively explained with the Gouy-Chapman theory, but with negatively charged ones deviations are observed. At a particular pH, the divalent ion Ca²⁺ strongly influences the intensity of the spectra in the presence of negatively charged liposomes, but has no effect with neutral liposomes.

With mitochondrial membranes an effect of Ca²⁺ similar to that with negatively charged liposomes is observed. Depletion of the phospholipids of the mitochondria and subsequent restoration of the mitochondrial membrane with lecithin, strongly diminishes this effect, but restoration with negatively charged phospholipids does not influence it.

From these observations it is concluded that the anionic form of the uncoupler molecule when bound to mitochondria is located within the partly negatively charged phospholipid moiety of the membrane, with its anionic group pointing to the aqueous solution.     

Induction of mutants in Saccharomyces cerevisiae by guanidine hydrochloride II. Conditions that prevent induction

The induction of ⁻ “petite” mutants by guanidine hydrochloride (GuHCl) is inhibited in several conditions. Anaerobiosis inhibited the induction either with or without cell multiplication. Both nalidixic acid (NA) and cycloheximide (CH) inhibited the induction of mutants. On the other hand, chloramphenicol (CAP) produced a dual effect: at low concentration it stimulated, at high concentration it inhibited, the induction. The effect of these different inhibitors on the transformation of ⁺ mother cells into ⁻ by GuHCl is discussed.     

Antifungal effect of CopA3 monomer peptide via membrane-active mechanism and stability to proteolysis of enantiomeric d-CopA3

In our previous study, coprisin, a 43-mer defensin-like peptide, was derived from the dung beetle, Copris tripartitus, and a 9-mer CopA3 (monomer), truncated coprisin analog peptide, was designed. However, the antifungal effects of CopA3 are not known yet. In this study, the antifungal activity and mechanism of CopA3 were investigated and to develop a more effective antimicrobial peptide under physiological conditions, the enantiomeric d-CopA3 was designed. l- and d-CopA3 had a similar antifungal activity without chiral selectivity, and their activity was more potent than that of melittin used as a positive control. Furthermore, l- and d-CopA3 did not even show any hemolysis against human erythrocytes. Membrane studies using propidium iodide and bis-(1,3-dibutylbarbituric acid) trimethine oxonol [DiBAC₄(3)], suggested that the antifungal effect of l- and d-CopA3 was due to the membrane-active mechanism, by contrast with coprisin possessing apoptotic mechanism without membrane permeabilization. Finally, the proteolytic resistance and antifungal activity of l- and d-CopA3 against trypsin was analyzed by HPLC and colony count assay. The results showed that only d-CopA3 maintained a potent antifungal activity despite the proteolytic condition. Therefore, this study suggests that d-CopA3 has potential as a novel antimicrobial agent.     

小儿肺热咳喘口服液联合三联吸入雾化治疗法对哮喘急性发作的影响

目的:探讨小儿肺热咳喘口服液联合三联吸入雾化治疗方案对哮喘患儿的治疗效果以及对肺功能的影响作用。方法:将我院自2017年1月至2018年11月间收治的哮喘患儿210例作为研究对象,按照随机数字表法分为两组各105例,研究组患儿在布地奈德、沙丁胺醇、异丙托溴铵三联吸入雾化治疗的基础上给予小儿肺热咳喘口服液进行治疗,对照组患儿仅给予三联雾化吸入治疗,对比观察两组患儿的疗效和预后。结果:研究组临床治疗后总有效率为95.24%,明显高于对照组77.14%(P0.05);治疗后研究组咳嗽消失时间、呼吸困难消失时间和急性发作随诊时间均明显短于对照组(P0.05),两组肺部喘鸣音消失时间比较差异无统计学意义(P0.05);治疗前患儿第一秒用力呼气量(forced expiratory volume in one second,FEV1)、最大肺活量(forced vital capacity,FVC)及FEV1/FVC值、呼气峰流速值(peak expiratory flowrate,PEF)对比无统计学意义(P0.05),治疗后1 d、3 d、7 d以上指标水平均明显升高,且在治疗后7 d,研究组明显高于对照组(P0.05)。治疗期间研究组有1例出现轻度腹泻,3例食欲减退,并发症的发生率为3.81%(4/105),对照组治疗期间2例出现轻度腹泻,4例食欲减退,并发症的发生率为5.71%(6/105),两组比较无统计学意义(P0.05)。结论:使用小儿肺热咳喘口服液联合三联吸入雾化法治疗儿童哮喘急性发作,可改善患儿临床症状和肺功能,疗效显著,可推广使用。     

布地奈德福莫特罗联合噻托溴铵吸入剂治疗慢阻肺疗效及对CRP、SaO2、PH水平影响

摘要 目的：探究布地奈德福莫特罗联合噻托溴铵吸入剂对慢阻肺治疗效果及对患者C-反应蛋白（C reaction protein,CRP）、血氧饱和度（arterial oxygen saturation,SaO₂）以及肺动脉高压（pulmonary Hypertension,PH）的影响。方法：选择2016年至2019年于我院接受治疗的82例慢性阻塞性肺病患者,按照随机数字表法将其均分为两组（各41例）,对照组单纯接受布地奈德福莫特罗治疗,研究组在对照组基础上加用噻托溴铵吸入剂施治,对比两组治疗有效率,治疗前后的第1秒用力呼气容积（forced expiratory volume in one second,FEV₁）、用力肺活量（forced vital capacity,FVC）、第1秒用力呼气容积占预计值百分比（Percentage of FEV₁ in the predicted value,FEV₁%）、二氧化碳分压（carbon dioxide partial pressure,PaCO₂）、SaO₂、PH值、CRP。结果：（1）研究组治疗有效率明显高于对照组（P<0.05）;（2）治疗前两组FEV₁、FVC、FEV₁%对比无差异（P>0.05）,治疗后研究组上述指标均高于对照组（P<0.05）;（3）治疗前两组SaO₂、PaCO₂、PH值对比无差异（P>0.05）,治疗后研究组SaO₂、PH值均高于对照组,PaCO₂低于对照组（P<0.05）;（4）治疗前两组CRP水平无差异（P>0.05）,治疗后研究组上述因子水平均低于对照组（P<0.05）。结论：布地奈德福莫特罗联合噻托溴铵吸入剂对慢阻肺具有较好的治疗效果,能够显著改善患者肺功能及血气指标,同时还能够缓解患者炎性状态。