Calcium-dependent protease of the cyanobacterium Anabaena: molecular cloning and expression of the gene in Escherichia coli, sequencing and site-directed mutagenesis

Summary It has been suggested that a calcium-dependent intracellular protease of the cyanobacterium, Anabaena sp., participates in the differentiation of heterocysts, cells that are specialized for fixation of N₂. Clones of the structural gene (designated prcA) for this protease from Anabaena variabilis strain ATCC 29413 and Anabaena sp. strain PCC 7120 were identified via their expression in Escherichia coli. The prcA gene from A. variabilis was sequenced. The genes of both strains, mutated by insertion of a drug resistance cassette, were returned to these same strains of Anabaena on suicide plasmids. The method of sacB-mediated positive selection for double recombinants was used to achieve replacement of the wild-type prcA genes by the mutated forms. The resulting mutants, which lacked Ca²⁺-dependent protease activity, were not impaired in heterocyst formation and grew on N₂ as sole nitrogen source.     

Immunocytochemical Localization of Glutamine Synthetase in Organs of Phaseolus vulgaris L

Glutamine synthetase was localized in nodules, roots, stems, and leaves of red kidney bean (Phaseolus vulgaris L.) by immunocytochemistry. Affinity purified antibodies reactive with glutamine synthetase were prepared using purified nodule-enhanced glutamine synthetase. Immunogold labeling was observed in the cell cytoplasm in each plant organ. In nodules, the labeling was more intense in the infected cells than in the uninfected cells. No labeling was observed in nodule bacteroids, peribacteroid spaces, or in peribacteroid membranes, while previous reports of glutamine synthetase immunolabeling of legume nodules showed labeling in the bacteroid fraction. Significant labeling was observed in nodule proplastids which contained starch granules. Substantial labeling was also observed in leaf chloroplasts. No labeling was observed in other organelles including mitochondria, peroxisomes, and endoplasmic reticulum. Preimmune IgGs did not bind to any structure in the tissues examined.     

A mechanistic perspective on bacterial metabolism of chlorinated methanes

Chlorinated methanes are important environmental pollutants, which can be metabolized by bacteria. The biotransformation of chlorinated methanes by bacteria has been shown to be due either to gratuitous metabolism (cometabolism) or their use as a source of carbon and energy. The reactions which result in carbon-halogen bond cleavage include substitutive, reductive, oxygenative, and gem-elimination mechanisms. Certain methylotrophic bacteria can use dichloromethane as a source of carbon and energy. Dichloromethane dehalogenase catalyzes the first substitutive reaction in this metabolism. The enzyme shows a 10¹⁰-fold rate enhancement over the reaction of the bisulfide anion with dichloromethane in water. Pseudomonas putida G786 synthesizes cytochrome P-450_CAM which catalyzes the gratuitous reduction of chlorinated methanes. These studies with purified enzymes are beginning to reveal more detailed mechanistic features of bacterial chlorinated methane metabolism.Abbreviations DNA deoxyribonucleic acid - k_cat catalytic first order rate constant for an enzyme catalyzed reaction - K_M Michaelis constant for an enzyme catalyzed reaction - MNDO modified neglect of diatomic overlap - PIMA pattern induced multialignment - DCMD dichloromethane dehalogenase     

Pollen-specific expression of theArabidopsis thaliana α1-tubulin promoter assayed by β-glucuronidase,chloramphenicol acetyltransferase and diphtheria toxin reporter genespromoter assayed by β-glucuronidase,chloramphenicol acetyltransferase and diphtheria toxin reporter genes

We have characterized the promoter specificity of theArabidopsis thaliana α₁-tubulin (α ₁-tub) gene by studying expression patterns of gene fusions between the 2.2 kbp 5′ upstream region of theα ₁-tub gene and each of three different reporters: chloramphenical acetyltransferase, β-glucuronidase or the diphtheria toxin chain A gene. Analysis of transgenic tobacco andArabidopsis plants carrying the transgene showed that the chloramphenicol acetyltransferase and β-glucuronidase activities were not detected in any vegetative or reproductive organs except mature pollen. Transgenic tobacco plants carrying the diphtheria toxin chain A gene under the control of theα ₁-tub promoter were of normal phenotype but seed fertility was drastically reduced. Furthermore, the transgene could not be transmitted to the next generation through pollen, supporting the observation that theα ₁-tub promoter is active only in pollen. It was observed that the promoter activity was most active in mature pollen and decreased significantly duringin vitro pollen germination, indicating that the promoter is inactive or subdued in germinating pollen. The promoter activity was not affected by various plant growth hormones during pollen maturation.     

脂肪族类两亲物对肌浆网膜脂区结构的影响

C18饱和脂肪酸和胺可增加DPH标记肌浆网(SR)的荧光偏振度,而C18单不饱和脂肪酸。胺和醇则使其偏振度下降。加入MgATP,可除去单不饱和脂肪胺引起的DPH标记的荧光偏振度下降,并使之高于未加脂肪胺的对照水平。饱和酸及相应胺可使标记于膜脂中层和深层的TAS和12AS的荧光偏振度上升,不饱和酸及相应胺和醇仅使12AS荧光偏振下降。说明脂肪族类两亲物对SR膜流动性的影响与脂肪链饱和程度有关。饱和者主要使膜中、深层流动性下降.不饱和者主要使膜深层流动性升高。     

青龙河底栖无脊椎动物群落结构及其水质评价

本文论述了青龙河底栖生物种类、数量、分布和结构等特点及其与环境因子间的关系。应用Beck、Gleason、Shannon、Simpson等生物指数对水质状况进行评价。结果表明青龙河除个别断面受污染外,大部分河段属尚清洁水。