Effect of papulacandin B on β-glucan synthesis in Schizosaccharomyces pombe

Abstract The antifungal antibiotic papulacandin β inhibited B(1,3)glucan-synthase activity, in vitro, from Schizosaccharomyces pombe . Levels of β(1,3)glucan-synthase from antibiotic-treated cultures were lower than the control cultures whereas mannan-synthase and β(1,3)glucanase activities were almost unaffected. The presence of an osmotic stabilizer reduced the inhibition of growth caused by the antibiotic. Addition of papulacandin β to a culture of S. pombe specifically inhibited incorporation of glucose into the β-glucan cell wall fraction. The fatty acids as well as the hydroxyl groups on the phenol residue of the papulacandin β molecule were essential for the inhibitory activity.     

Mutants of Saccharomyces cerevisiae cell division cycle defective in cytokinesis. Biosynthesis of the cell wall and morphology

The four temperature-sensitive mutants of Saccharomyces cerevisiae in the cell division cycle defective in cytokinesis (cdc, 3, 10, 11 and 12), have been analyzed with respect to the biosynthesis of the cell wall polymers. After 3 hours of incubation at the non-permissive temperature (37°C) these strains stop growing. The synthesis of glucan, mannan and chitin (wall polymers) level off in a similar time, but glucan, mannan and chitin synthases remained active for at least 4 hours.If the mutants are analyzed by transmission and scanning electron microscopy different pictures emerge. Two of the mutants cdc 10 and cdc 12, after 3 hours of incubation at 37°C present apparently normal cytoplasms and cell wall surfaces with multiple elongated buds. The other two mutants, cdc 3 and cdc 11, present a completely disarranged cytoplasmic content and damage at the level of the plasma membrane is evident.These and other observations, suggest that between the execution points of cdc 3 (0.27) and cdc 10 (0.58), essential processes in the assembly of cell membrane occur.This work was supported in part by a grant from la Comisión de Investigación Científica y Técnica of the Spanish Ministerio de Educación y Ciencia (Project no. 4593-1980).     

The product of the nitrogen fixation regulatory gene nfrX of Azotobacter vinelandii is functionally and structurally homologous to the uridylyltransferase encoded by glnD in enteric bacteria.

We sequenced the nitrogen fixation regulatory gene nfrX from Azotobacter vinelandii, mutations in which cause a Nif- phenotype, and found that it encodes a 105-kDa protein (NfrX), the N terminus of which is highly homologous to that of the uridylyltransferase-uridylyl-removing enzyme encoded by glnD in Escherichia coli. In vivo complementation experiments demonstrate that the glnD and nfrX products are functionally interchangeable. A vinelandii nfrX thus appears to encode a uridylyltransferase-uridylyl-removing enzyme, and in this paper we report the first sequence of such a protein. The Nif- phenotype of nfrX mutants can be suppressed by a second mutation in a recently identified nifL-like gene immediately upstream of nifA in A. vinelandii. NifL mediates nif regulation in response to the N status in A. vinelandii, presumably by inhibiting NifA activator function as occurs in Klebsiella pneumoniae; thus, one role of NfrX is to modify, either directly or indirectly, the activity of the nifL product.     

The granin (chromogranin/secretogranin) family

The chromogranins/secretogranins, referred to in abbreviated form as granins, are a family of acidic secretory proteins that are found in the secretory granules of a wide variety of endocrine cells and neurons, being stored together with many different peptide hormones and neuropeptides. The recent elucidation of their primary structure has provided insights into possible functions of these proteins. Moreover, the granins have been successfully used as markers for normal and neoplastic endocrine and neuronal cells, as well as model proteins to understand the sorting mechanism involved in the formation of secretory granules.     

Approach to the analysis of diuretics and masking agents by high-performance liquid chromatography—mass spectrometry in doping control

The application of thermospray and plasmaspray high-performance liquid chromatography—mass spectrometry to the analysis of diuretics and probenecid has been investigated. The latter method gave better ionization efficiency than the former, and its response was optimized by altering the solvent composition: best results were obtained with water—methanol—acetonitrile—trifluoroacetic acid. Using different proportions of these solvents, three isocratic systems were developed to separate the compounds under study. The principal characteristic of plasmaspray positive-ion mass spectra was a protonated molecular ion and very little fragmentation was evident. In the negative ionization mode, the plasmaspray method gave mass spectra showing more fragmentation, which resulted in additional structural information. The ability of trifluoroacetic acid to form negative cluster ions precluded its use as a mobile phase component. The minimum detectable amounts determined by the analysis in the positive-ion mode was compound-dependent, but generally ca. 10–150 ng. In many cases the compounds could be detected in urine extracts.     

Redescription of Branchinecta granulosa Daday, 1902 from Argentina (Crustacea: Branchiopoda)

The species Branchinecta granulosa Daday 1902, is redescribed on the basis of adult material from near Facundo (Chubut Province, Argentina). Its relationship to its regional congeners is outlined.     

Calcium binding to the cytosol and calcium extrusion mechanisms in intact synaptosomes and their alterations with aging.

A simple method to measure cytosolic calcium binding in intact presynaptic nerve terminals (synaptosomes) from rat brain, which is based on the simultaneous determination of [Ca2+]i and total [45Ca2+] in quin2-loaded synaptosomes undergoing a switch from high- to low-calcium containing medium, is presented. Binding to the cytosolic compartment alone was obtained following depletion of calcium storing organelles in the presence of carbonyl cyanide p-trifluoromethoxyphenylhydrazone/oligomycin plus caffeine. Synaptosomes, as compared to various cells types, have a high calcium binding capacity to the cytosolic compartment; maximum binding, Ca.Bmax, was 4.76 mM and calculated s0.5 was 218 nM. Calcium binding to the cytosolic compartment as a function of aging was also determined; Ca.Bmax was reduced to 1.84 mM and s0.5 increased to 492 nM in 30-month-old rats, indicating that the buffering of high calcium loads is impaired in old animals. The results obtained for binding of calcium to mitochondria and caffeine-sensitive calcium stores are consistent with an age-dependent reduction in calcium bound to mitochondria, whereas caffeine-sensitive calcium stores were unaffected. Finally, we have estimated the net rates of calcium extrusion in intact synaptosomes, and found that calcium efflux through the Na/Ca exchanger and Ca(2+)-ATPase was markedly reduced in old rats.     

Biological characterization of Trichinella isolates from various host species and geographical regions.

Forty isolates of Trichinella collected from 5 continents were compared for 7 biological characters: newborn larvae produced per female worm cultured in vitro at the seventh, eighth, and ninth day postinfection, host muscle nurse cell development time, reproductive capacity index in rats and chickens, and resistance of muscle larvae to freezing. The isolates also were compared by analyses of an environmental character of the location from which they were isolated: the isotherms for January and July. By factorial analysis of correspondence of the biological and environmental data, the 40 isolates were grouped into 8 gene pools (T1-T8). The environmental temperature-related distribution was more evident for the sylvatic isolates (T2, T3, T5, T6, T7, T8), than for T1, which was isolated from domestic pigs, and for T4, a bird-adapted, nonencapsulating genetic type. The 8 biological groups correlated closely with the 8 gene pools previously identified on the basis of allozyme analysis. These results support the concept that the genus Trichinella is composed of at least 5 distinct gene pools or sibling species: Trichinella spiralis sensu stricto (T1), Trichinella nativa (T2), Trichinella sp. (T3), Trichinella pseudospiralis (T4), and Trichinella nelsoni (T7), and 3 other groups of uncertain taxonomic status (i.e., T5, T6, and T8).     

Distance measurements using paramagnetic ion-induced relaxation in the saturation transfer electron spin resonance of spin-labeled biomolecules: Application to phospholipid bilayers and interdigitated gel phases

The saturation transfer electron spin resonance (STESR) spectra of spin-labeled phosphatidylcholines in gel phase lipid bilayers are shown to be sensitive to dipolar spin-spin interactions with paramagnetic ions in the aqueous phase. The reciprocal integrated intensity of the STESR spectrum is linearly dependent on aqueous Ni²⁺ ion concentration, hence, confirming the expectation that the STESR intensity is directly proportional to the spin-lattice relaxation time of the spin label. The gradient of the relaxation rate with respect to Ni²⁺ ion concentration decreases strongly with the position of the nitroxide group down the sn-2 chain of the spin-labeled lipid and is consistent with a 1/R³ dependence on the distance, R, from the bilayer surface. The values derived for the dimensions of the bilayer and lipid molecules in the case of dipalmitoyl phosphatidylcholine (DPPC) are in good agreement with those available from x-ray diffraction studies. Allowance for the multibilayer nature of the DPPC dispersions gives an estimate of the water layer thickness that is also consistent with results from x-ray diffraction. The profile of the paramagnetic ion-induced relaxation is drastically changed with DPPC dispersions in glycerol for which the lipid chains are known to be interdigitated in the gel phase. The terminal methyl groups of the lipid chains are located approximately in register with the C-3 atoms of the sn-2 chain of the oppositely oriented lipid molecules in the interdigitated phase. The thickness of the lipid layer and the effective thickness of the lipid polar group are reduced by ~40% in the interdigitated phase as compared with the bilayer phase. The calibrations of the distance dependence established by use of spin labels at defined chain positions should be applicable to STESR measurements on other biological systems.