Purification and characterisation of an inducible β-galactosidase from Corynebacterium murisepticum

The -galactosidase (EC 3.2.1.32) of Corynebacterium murisepticum (inducible by lactose and galactose) was purified by successive column chromatography on Sephadex G-200, DEAE-Sephadex A-50 and DEAE-cellulose (DE52). The enzyme was found to be a dimer of identical subunits of molecular mass 100,000 daltons. The K _m values of the enzyme for the substrates lactose and o-nitrophenyl--d-galactopyranoside (ONPG) are 16.7 mM and 4.4 mM, respectively, indicating, its low affinity for the substrates. The Ouchterlony immunodiffusion method exhibited immunological homogeneity of the enzyme preparation. The catalytic site of the enzyme does not take part in antigen-antibody reaction.     

The role of tungstate and/or molybdate in the formation of aldehyde oxidoreductase in Clostridium thermoaceticum and other acetogens; immunological distances of such enzymes

Besides Clostridium thermoaceticum and C. formicoaceticum other resting acetogenic clostridia such as C. aceticum and C. thermoautotrophicum and to a lesser extent non-clostridial acetogens such as Butyribacterium methylotrophicum and Eubacterium limosum were able to reduce propionate to propanol at the expense of carbon monoxide or formate. Methylviologen usually increased the reduction rate. Ten M molybdate in the growth medium decreased this capability for C. thermoaceticum but increased it or had no effect for the other organisms. Ten M tungstate in the growth medium increased the aldehyde oxidoreductase activity in all organisms. Crude extracts of C. thermoaceticum cells grown in the presence of 10 M or 1 mM molybdate showed by ELISA the same or even a 4 fold concentration of aldehyde oxidoreductase in the latter case. However, the enzymic activity was very low in both cases. Omission of dithionite in the growth medium diminished the antigen by a factor of about 8. The immunological distance between the enzyme from C. thermoaceticum and C. thermoautotrophicum was rather low but very large to C. formicoaceticum and undeterminably large to the other organisms.Abbreviations Ald-DH aldehyde dehydrogenase - AOR aldehyde oxidoreductase - CO-DH carbon-monoxide dehydrogenase - ELI-SA enzyme-linked immunosorbent assay - FDH formate dehydrogenase - MV methylviologen - V⁺⁺ oxidised - V^+. reduced viologen     

A Kdp-like,high-affinity,K+-translocating ATPase is expressed during growth of Rhodobacter sphaeroides in low potassium media

Cells of the purple non-sulphur bacterium Rhodobacter sphaeroides express a high-affinity K⁺ uptake system when grown in media with low K⁺ concentrations. Antibodies againts the catalytic KdpB protein or the whole KdpABC complex of Escherichia coli crossreact with a 70.0 kDa R. sphaeroides protein that was expressed only in cells grown in media with low K⁺ concentrations. In membranes derived from R. sphaeroides cells grown with low K⁺ concentrations (induced cells), a high ATPase activity could be detected when assayed in Tris-HCl pH 8.0 containing 1 mM MgSO₄. This ATPase activity increased upon addition of 1 mM KCl from 166 to 289 mol ATP hydrolysed x min^-1 x g protein^-1 (1.7-fold stimulation). The K⁺-stimulated ATPase activity was inhibited approximately 93% by 0.5 mM vanadate but hardly by N,N-dicyclohexylcarbo-diimide (DCCD). These results indicate that the inducible K⁺-ATPase in R. sphaeroides resembles the Kdp K⁺-translocating ATPase of Escherichia coli. This Kdp-like transport system is also expressed in R. capsulatus and Rhodospirillum rubrum during growth in media with low K⁺ concentrations suggesting a wide distribution of this transport system among phototrophic bacteria.Abbreviations electrical potential difference across the cytoplasmic membrane - pH pH difference across the cytoplasmic membrane - BSA bovine serum albumine - PAGE polyacrylamide gel electrophoresis - HEPES 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid - PMSF phenyl-methyl-sulfonyl fluoride - DCCD N,N-dicyclohexylcarbodiimide - AIB 2--aminoisobutyric acid - TMG methyl--d-thiogalactopyranoside     

A 43 kDa protein inserted at the plasma membrane-cytoskeleton interface in rumen entodiniomorphid ciliates

Summary The P-43 ofEudiplodinium and homologous proteins in three other entodiniomorphid species, free-living ciliates, flagellates, and HeLa cells, were identified at the plasma membrane-cytoskeleton interface. Proteins cross-reacting with MAb B6 were also located at the ciliary inner surface of the plasma membrane. Due to the strong adhesion of the plasma membrane to the underlying cytoskeleton, classical extraction with detergents, urea, NaOH, and PTA, failed to separate the two components completely. However, the extraction properties of P-43, associated with its membrane-cytoskeleton interactive functions, suggest that this unglycosylated protein may present some analogies with proteins of the intermediate filaments. Their ubiquity and localization suggest that P-43 and MAb B6 crossreacting proteins may not be strictly epiplasmic but could be amphitropic proteins, strongly anchored to both the plasma membrane and the underlying microfilament framework, via protein-protein binding or by direct insertion in the lipid bilayer.Abbreviations BSA bovine serum albumin - Con A concanavalin A - EDTA ethylene diamine tetraacetic acid - EM electron microscopy - IF intermediate filaments - MAb monoclonal antibody - MET 2-mercaptoethanol - MW molecular weight - PAb polyclonal antibody - PBS phosphate buffered saline - PMSF phenylmethylsulfonyl fluoride - PTA phosphotungstic acid - SDS sodiumdodecyl sulfate - TAME Na-p-tosyl-arginine methyl ester - TLCK Na-p-tosyl-lysine chloromethyl ketone     

Immunological determination of digestive rates in the syntopic scorpions Urodacus armatus Pocock and Urodacus novaehollandiae Peters

Scorpions have lengthy periods of inactivity, which may be caused either by a long physiological digestion time or predator avoidance or both. We used a quantitative immunological assay to monitor the amount of prey antigen remaining in the hepatopancreas of Urodacus armatus and U. novaehollandiae for 32 days after a meal to test the hypothesis of a long digestion time. In both species, prey antigen concentration in the hepatopancreas increased between 0.25 and 0.5 days after feeding, probably because of incomplete transfer of prey proteins from the large preoral cavity to the hepatopancreas at this stage of digestion. This was followed by a rapid decline, and by 3 days after feeding less than 25% of ingested antigen remained in each species. Small prey residues could still be detected up to 32 days. Prey decay curves in both species were best fitted by a log-log transformation, and were not significantly different. Since digestion is essentially complete after 3 days, it appears that longer periods of post-feeding inactivity are for predator avoidance, not digestion. However, the 32-day detection period exceeds those reported for other invertebrates, and suggests an unusual digestive physiology in scorptions.     

高等植物叶绿体和线粒体免疫亲近性的研究

以火箭免疫电泳分析表明:大豆叶绿体抗体与大豆线粒体有免疫交叉反应,同时大豆线粒体抗体与大豆叶绿体也有免疫交叉反应,但是大豆线粒体的抗体与鼠肝线粒体之间无免疫交叉反应。这说明高等植物线粒体对叶绿体比之对动物线粒体在免疫特性上有更大的亲近性,亦即高等植物线粒体和高等植物的叶绿体有更大的同源性。经火箭免疫电泳、交叉免疫电泳和线状免疫电泳进一步分析表明:菠菜偶联因子抗体(AbCF_1)和大豆线粒体、大豆叶绿体间,大豆线粒体抗体与CF_1和大豆叶绿体之间,以及大豆叶绿体的抗体(AbC)与CF_1和大豆线粒体间有免疫交叉反应,说明两种换能器之间有免疫亲近性,并分别与CF_1存在免疫亲近性。这揭示两种换能器免疫亲近性的表现是由于存在共同物质基础所致,这内在共同物质基础是偶联因子。这个结果有力地支持高等植物叶绿体和线粒体在结构和功能上以及发生上存在同源性的观点,在理论上也为两种换能器的起源和演化上存在同源性提供了一些依据。     

Lipoprotein immunogenetics in primates. I. Two serum beta-lipoprotein allotypes (Lmb1 and Lmb11) in rhesus monkeys and the LP-B immunological relationship with other primates

Immunogenetic investigations on two serum beta-lipoprotein allotypes of rhesus monkeys (Macaca mulatta) are reported. The allotypes, designated Lmb1 and Lmb11, are associated with the main lipoprotein family, LP-B or beta-lipoprotein, expressed on independent beta-molecules, and classify rhesus monkeys into three phenotypes: Lmb1, Lmb11, and Lmb1,11. Genetic and molecular studies indicate that the allotypes are encoded by two codominant autosomal allelic genes, Lmb1 and Lmb11. Anti-Lmb1 cross-reacts with the sera of two other macaque species, whereas anti-Lmb11 with sera of all Old World monkeys. Heteroimmune sera, antihuman apo-B and antirhesus LP-B, showed high but diversified degrees of cross reactivity with other primates.     

Characterization and Purification from Human Brain of a Hyaluronic Acid-Binding Glycoprotein, Hyaluronectin

Using affinity chromatography and enzyme-labelled immunological assays combined with affinity adsorption, we have obtained evidence for the binding of a brain glycoprotein to hyaluronic acid, and on this basis named it hyaluronectin. This binding was inhibited by hyaluronic acid and by the products of its hydrolysis by hyaluronidase from bovine testis, but was not inhibited by other glycosaminoglycans or by monosaccharides. Preparative affinity chromatography of brain acid-soluble proteins produced hyaluronectin in a good degree of purity. Contamination by albumin was less than 1% and the yield was as high as 80%.     

Territorial damselflies do not show immunological priming in the wild

Adaptive immunity allows vertebrates to gain protection against repeated pathogenic infections. Analogous responses (priming) have been recently uncovered in invertebrates. However, whether such responses are widespread is not known. The present study investigated the presence of immunological priming in males of a species whose phylogenetic position places it in one of the less derived insect orders. It is hypothesized that the efficiency of such a response could be related to animal condition, as assessed by the expression of a sexually selected ornament. Hetaerina americana Fabricius (Odonata: Calopterygidae) males bear a conspicuous ornament (a red wing spot), which is evolutionarily maintained via male territorial competition. Using field‐collected animals, a group of males is challenged with bacteria before exposure to a higher dose of the same or a different bacteria, and survival is compared with that of infected males not previously challenged, as well as control groups. Gram‐positive and Gram‐negative bacteria are used. To explore how long priming may take to work, the second exposure is carried out either after 1 or 5 days. Red spot and body size are entered in the analysis as predictors of survival within and between groups. There is no difference in survival among groups, which suggests no priming effect. Overall, red spot and body size are not consistent in explaining survival.     

盐酸右美托咪定联合盐酸罗哌卡因胸椎旁神经阻滞对肺癌根治术患者血清炎性因子和免疫学指标的影响

摘要 目的：探讨盐酸右美托咪定联合盐酸罗哌卡因胸椎旁神经阻滞（TPVB）对肺癌根治术患者血清炎性因子和免疫学指标的影响。方法：选取2018年4月~2020年4月期间于我院行肺癌根治术的患者400例,根据信封抽签法分为对照组和观察组,各200例,对照组给予盐酸罗哌卡因TPVB,观察组在对照组基础上联合盐酸右美托咪定,对比两组疼痛、炎性因子、免疫学指标、血流动力学及不良反应。结果：观察组术后6 h（T5）~术后48h（T8）时间点视觉模拟评分法（VAS）评分均低于对照组（P<0.05）。对照组插管后5 min（T1）~术毕（T3）时间点心率（HR）及平均动脉压（MAP）较麻醉诱导前（T0）时间点升高（P<0.013）,观察组T1~T3时间点HR、MAP低于对照组（P<0.05）。两组T8时间点白介素-6（IL-6）、C反应蛋白（CRP）、肿瘤坏死因子-α（TNF-α）均较T0下降,且观察组低于对照组（P<0.05）。两组T8时间点CD4⁺、CD4⁺/CD8⁺均较T0时间点下降,但观察组高于对照组（P<0.05）,CD8⁺均较T0时间点升高,但观察组低于对照组（P<0.05）。观察组不良反应总发生率低于对照组（P<0.05）。结论：盐酸右美托咪定联合盐酸罗哌卡因TPVB用于肺癌根治术患者,可稳定血流动力学,且可获得较好的镇痛效果,降低不良反应发生率,减轻术后炎性损伤及免疫抑制。