Schistosoma mansoni: Inhibition of glucosephosphate isomerase and glycolysis by sugar phosphates

Glucosephosphate isomerase (EC 5.3.1.9) of Schistosoma mansoni is inhibited competitively by a number of tetrose, pentose, and hexose phosphates with inhibitor constant (K_i) values in the range of 0.5 to 400 μM. The most potent inhibitor is 5-phospho-d-arabinonate which resembles the cis-enediolate transition state intermediate of the reaction. These analogs were also found to be effective inhibitors of the production of lactate from glucose by suitably supplemented worm homogenates. The rank order of potency of inhibition of glycolysis was inversely related to the magnitudes of the K_i values for glucosephosphate isomerase. These K_i values were similar to those previously reported for mammalian glucosephosphate isomerase, suggesting similarities in the steric and electronic characteristics of the active sites of these isofunctional enzymes. This conclusion was further supported by the observed pH dependence of the inhibition by 5-phospho-d-arabinonate. Although glucosephosphate isomerase is not a rate-limiting enzyme of glycolysis, in the conventional sense, its selective inhibition could be of chemotherapeutic importance, in part because of the accumulation in glycolyzing systems of glucose 6-phosphate which is a potent feedback inhibitor of hexokinase.     

The use of phenylmethylsulfonyl fluoride in the study of catabolite inactivation and repression in intact cells of Saccharomyces cerevisiae

Catabolite inactivation of fructose-1,6-bisphosphatase, isocitrate lyase, phosphoenolpruvate carboxykinase and malate dehydrogenase in intact cells could be prevented by phenylmethylsulfonyl fluoride added 40 min prior to the addition of glucose. Protein synthesis, fermentative and respiratory activity and catabolite repression were not affected. Elimination of catabolite inactivation by the addition of PMSF revealed that catabolite repression started at different times for different enzyme.Abbreviation PMSF phenylmethylsulfonyl fluoride     

Parameters of unbalanced growth and reversible inhibition of deoxyribonucleic acid synthesis in Brevibacterium ammoniagenes ATCC 6872 induced by depletion of Mn2+. Inhibitor studies on the reversibility of deoxyribonucleic acid synthesis

Unbalanced growth induced by depletion of manganese ions was a prerequisite for production of ribonucleotides in a high salt mineral medium with the wildtype strain Brevibacterium ammoniagenes ATCC 6872. The concentration of manganese strictly controlled the overall deoxyribonucleic acid (DNA) synthesis, whereas ribonucleic acid (RNA), protein and cell wall synthesis remained essentially unimpaired in the manganese-lacking cells.The reversibility of inhibition of overall DNA synthesis was shown by enhanced incorporation (up to threefold compared to the cultures supplied with sufficient manganese) of [8-¹⁴C] adenine into alkali-stable, trichloroacetic acid-insoluble material after subsequent addition of 10 M MnCl₂ to 15 h-old depleted cultures.The results of inhibitor studies on the restoration of overall DNA synthesis due to subsequent addition of manganese ions to depleted cultures suggest that ribonucleotide reduction is the primary target of the manganese starvation during nucleotide fermentation with Brevibacterium ammoniagenes ATCC 6872.     

In vitro degradation of guanosine 3′,5′-bis(diphosphate) [ppGpp] by the spoT gene product [ppGppase] from auxotrophic strains of Escherichia coli: Effects of various antibiotics and drugs

The enzyme specifically hydrolyzing guanosine 3,5-bis(diphosphate) [ppGpp] has been isolated from the ribosomal fraction of Escherichia coli; it released pyrophosphate from the 3-position of ppGpp. The effects of various drugs and antibiotics known to interfere with protein and/or RNA synthesis were investigated in the ppGpp degrading reaction. It was determined that tetracycline, chlorotetracycline, and thiostrepton strongly inhibited the reaction, whereas levallorphan gave a moderate inhibition. Only the tetracycline-mediated inhibition could be reversed by manganese ions. Oxytetracycline, rifampicin, fusidic acid, kirromycin, streptomycin, puromycin, chloramphenicol, and morphine did not inhibit the decay reaction.Abbreviations ppGpp guanosine 3,5-bis(diphosphate)     

Adenosine deaminase from Azotobacter vinelandii. Purification and properties

Adenosine deaminase (EC 3.5.4.4) was found to occur in the extract of Azotobacter vinelandii, strain 0, and purified by heating at 65°C, fractionation with ammonium sulfate, DEAE-cellulose chromatography and gel filtration on Sephadex G-150. Purified adenosine deaminase was effectively stabilized by the addition of ethylene glycol. The molecular weight of the enzyme was estimated to be 66,000 by gel filtration on Sephadex G-150. The enzyme specifically attacked adenosine and 2-deoxyadenosine to the same extent, and formycin A to a lesser extent. The pH optimum of the enzyme was observed at pH 7.2. Double reciprocal plot of initial velocity versus adenosine concentration was concave upward, and Hill interaction coefficient was calculated to be 1.5, suggesting the allosteric binding of the substrate. ATP inhibited adenosine deaminase in an allosteric manner, whereas other nucleotides were without effect. The physiological significance of the enzyme was discussed in relation to salvage pathway of purine nucleotides.     

Inhibition of Ouabain-binding to (Na+ + K+)ATPase by antibody against the catalytic subunit but not by antibody against the glycoprotein subunit

Antibodies against purified ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)}\text{ATPase}$ from the rectal gland of Squalus acanthias, as well as against its catalytic subunit, inhibited ouabain binding by as much as 50%. However, antibodies against the glycoprotein subunit did not inhibit ouabain binding. These data suggest that binding of antibody against the catalytic subunit to the enzyme either covers the ouabain binding site or destroys its conformation, while binding of antibody against the glycoprotein has no such effect.     

The acetylene reduction technique as an assay for nitrogenase activity in the methane oxidizing bacterium Methylococcus capsulatus strain bath

The use of acetylene as a convenient assay substrate for nitrogenase in methane oxidising bacteria is complicated by the observation that it is a potent inhibitor of the methane monooxygenase enzyme in both whole cells and cell-free extracts. If the cells were provided with alternative oxidisable carbon substrates other than methane then nitrogen fixing cells would reduce acetylene to ethylene. Hydrogen gas also served as an oxidisable substrate in the assay. Nitrous oxide, which is reduced by nitrogenase to N₂ and H₂O, was not an inhibitor of methane monooxygenase function and could be used as a convenient assay substrate for nitrogenase. Reduction of both substrates by whole cells showed similar response to oxygen in the assay system and in this respect Methylococcus resembles other free living nitrogen fixing aerobes.     

对叶榕榕果内生细菌分离鉴定及抑菌活性的研究

【目的】为植物-内生细菌的生态关系研究, 以及植物内生细菌资源的利用提供一定的依据。【方法】采用微生物学传统分离培养的方法从对叶榕果实中分离到内生细菌54株, 通过限制性酶切分析(ARDRA)共有16个操作分类单元(Operational taxonomic units, OTUs), 对16株代表菌株16S rDNA序列系统发育分析。【结果】16个菌株都找到了与其相似性最高的菌株, 相似性达到95%?100%。其中6株为芽孢杆菌Bacillus属, 为对叶榕果实内生细菌优势菌属; 3株为Staphylococcus属, 2株为Pseudomonas属, 1株为Serrata属, 1株为非培养细菌的同源菌, 1株为Kocuria属, 1株为Delftia属, 还有1株为Acinetobacter属。【结论】这16株内生菌在系统发育树中明显聚为两大支; 在参与抑菌试验的14株内生菌中, 有13株对受试菌有不同程度的拮抗作用。尤其是其中的芽孢菌属的Swx15和不动杆菌属的Swx25菌株, 抑菌作用较强, 且有较广的抑菌谱性。     

假单胞菌YL11对扩展青霉的抑制作用及其机理初探

【背景】苹果青霉病是由扩展青霉引起的一种重要的果实采后病害,影响果实品质导致苹果腐烂从而造成经济损失。【目的】研究假单胞菌YL11对扩展青霉的抑制作用和苹果采后青霉病的防治效果,并对抑菌机理进行初步探讨。【方法】以扩展青霉为供试菌株,研究不同浓度的假单胞菌YL11无菌发酵液对扩展青霉菌落直径、孢子萌发率、菌丝体干重、苹果损伤接种病斑直径扩展的影响,利用对电导率、核酸及蛋白释放量、AKP含量、SDH活性、ATP酶活性和ATP含量的影响对抑菌机理进行探究。【结果】假单胞菌YL11无菌发酵液能有效抑制扩展青霉生长,抑菌圈直径为22.33±0.27 mm,抑菌效价为71.67 mm/mL;能有效抑制孢子萌发,100%无菌发酵液对孢子萌发抑制率达到80.2%;对扩展青霉的生物量也有一定抑制作用,体积分数为100%时,菌丝体干重为4.7mg/mL,抑制率达到39.74%;无菌发酵液处理能有效抑制苹果青霉病病斑的扩展,3d时对病斑扩展的抑制率最大,达到47.1%;无菌发酵液处理均能引起电导率升高、胞内核酸和蛋白释放量增大、胞外AKP含量升高、SDH活性降低、ATP酶活性和ATP含量均降低,且随着发酵液浓度的增加效果越明显。【结论】假单胞菌YL11能显著抑制扩展青霉的生长,破坏细胞膜结构、降低能量代谢酶活性,从而扰乱扩展青霉的正常生长,对苹果青霉病有较好的生防效果,具有潜在的开发价值。     

两株香蕉枯萎病拮抗细菌的筛选及抑菌机理

【目的】从发病蕉园中的健康香蕉根际筛选能有效抑制香蕉枯萎病病原菌生长的拮抗菌,进一步研究其抑菌机理。【方法】应用双层平板初筛和平板对峙实验复筛具有抑菌效果的拮抗菌;经生理生化试验、16S rRNA基因测序和特异引物扩增对拮抗菌进行鉴定;酸沉淀方法提取拮抗菌发酵液抑菌物质粗提液,基于比色法和HPLC测定粗提液对菌丝蛋白质含量、脂质过氧化、麦角甾醇和果胶酶活性的影响。【结果】筛选获得两株拮抗细菌H-2和H-7,初步鉴定为枯草芽孢杆菌和解淀粉芽孢杆菌,Gen Bank登录号分别为KX791428和KX791430;温室盆栽试验显示,两株拮抗菌对香蕉枯萎病的生防效率分别为59.1%和53.0%;H-2和H-7粗提液处理病原菌菌丝后,因脂质过氧化产生的丙二醛含量显著增加,分别达0.55μmol/L和0.48μmol/L;而蛋白含量、麦角甾醇含量和果胶酶活性均显著下降,其中H-2处理的抑制幅度更大,三项指标分别为0.15 mg/g、1.31 mg/g和0.008 7 U/m L,显著低于对照的0.25 mg/g、1.96 mg/g和0.035 U/m L。【结论】从健康香蕉根际筛选到两株拮抗细菌,两者可能通过增强病原菌菌丝的脂质过氧化和降低细胞代谢产物合成的方式抑制病原菌生长,可为两株拮抗菌的生防应用提供理论依据。