The isolation of protoplasts of the fission yeastSchizosaccharomyces byTrichoderma viride and snail enzymes

The formation of protoplasts of the fission yeastsSchizosaccharomyces pombe andSchizosaccharomyces versatilis after the combined application of snail enzymes andTrichoderma viride enzymes in an osmotic stabilizer (0.4m KC1, pH 5.5) was studied by light and electron microscopy. The effect of the enzymes used leads during 30 min to the formation of 100% protoplast population. Using electron microscopy no original walls or wall remnants were detected in the suspension of protoplasts. Protoplasts are viable and in liquid nutrient medium they regenerate cell walls and revert into normal cells. Such a protoplast population may be useful for biochemical study of protoplast metabolism by quantitative methods as well as for the chemical study of regenerating cell walls.     

Isoenzyme pattern of malate dehydrogenase during respiratory derepression in Schizosaccharomyces pombe

One isoenzyme of malate dehydrogenase with an isoelectric point of 6.4 was found in glucose-repressed cells of Schizosaccharomyces pombe. During respiratory derepression the activity of this isoenzymes decreased rapidly in vivo. In the course of this inactivation two new forms of malate dehydrogenase with isoelectric points of 6.0 and 5.7 appeared. It has been found that these two enzymic forms disappeared 4 h after the exhaustion of glucose; probably they are degradation products of the isoenzyme present in glucose-repressed cells. Fully derepressed cell of this fission yeast contain one isoenzyme of malate dehydrogenase with an isoelectric point of 5.3. The synthesis of this isoenzyme is initiated at glucose concentrations below 1.5 g/l.     

The Indigo of Commerce in Colonial North America

It can be demonstrated that the Indigo of Commerce in Colonial North America consisted of three species in the genusIndigofera. One of these was a native plant,I. Caroliniana Mill, while the other two were introduced.Indigofera tinctoria L. (French Indigo), an Old World species, andI. Suffruticosa Mill. (Guatemala Indigo), a New World species, were both introduced into South Carolina in the late 17th and early 18th centuries. Their cultivation flourished until the American Revolution. Neither of the introduced species has become naturalized in the Carolinas.     

Genetic variation in the activity of the histidine catabolic enzymes between inbred strains of mice: A structural locus for a cytosol histidine aminotransferase isozyme (Hat-1)

Variation in activity of the main histidine catabolic enzymes (histidase, urocanase, and aminotransferase) has been surveyed using inbred strains of mice (C57BL, DBA, Peru, SM, and SWR). Some variation was found in the activity of all enzymes, but only in the case of cytosolic histidine aminotransferase was it greater than twofold (SM 3.3-fold greater than C57BL). The divergent strains for the activity of this enzyme were crossed and the F ₁ 's were backcrossed; the segregation analysis indicated a single locus with additively acting alleles (designated Hat-1: a allele SM, b allele C57BL). Cytosolic histidine aminotransferase differed in heat stability between SM and C57BL, indicating that Hat-1 is a structural locus. The conflict in the biochemical literature (Morris et al., 1973; Noguchi et al., 1976a, b) over the number and subcellular distribution of the histidine aminotransferase isozymes is partly resolved by the acquisition of a variant at the Hat-1 locus. Hat-1 affects the cytosolic form but not the mitochondrial form of the enzyme. Purification and analysis of the isozymes of histidine aminotransferase from livers of C57BL and SM mice will further clarify the situation.     

A protective role of methionine-R-sulfoxide reductase against cadmium in Schizosaccharomyces pombe

The Schizosaccharomyces pombe cells harboring the methionine- R-sulfoxide reductase (MsrB)-overexpressing recombinant plasmid pFMetSO exhibited better growth than vector control cells, when shifted into fresh medium containing cadmium chloride (abbreviated as Cd). Although both groups of cells contained enhanced reactive oxygen species (ROS) and nitric oxide (NO) levels in the presence of Cd, ROS and NO levels were significantly lower in the S. pombe cells harboring pFMetSO than in vector control cells. Conversely, the S. pombe cells harboring pFMetSO possessed higher total glutathione (GSH) levels and a greater reduced/oxidized GSH ratio than vector control cells under the same conditions.     

Steroid derivatives

It was found in a study of the hydroxylating capacity of one of theFungi imperfecti, the genusBeauveria, some of the species of which are able to introduce the oxygen function into position 11α of various steroid compounds, that the enzyme activity ofBeauveria globulifera gives rise both to the corresponding 11α-hydroxy-derivative and to the 5β-saturated 11α-hydroxy-compound or a 5β-saturated 7β,11α-dihydroxy-compound. The isolation and the identification of the products of biotransformation of some steroids of the pregnane and androstane series byBeauveria globulifera is described.     

Structural and functional analysis of the anti-malarial drug target prolyl-tRNA synthetase

Aminoacyl-tRNA synthetases (aaRSs) drive protein translation in cells and hence these are essential enzymes across life. Inhibition of these enzymes can halt growth of an organism by stalling protein translation. Therefore, small molecule targeting of aaRS active sites is an attractive avenue from the perspective of developing anti-infectives. Febrifugine and its derivatives like halofuginone (HF) are known to inhibit prolyl-tRNA synthetase of malaria parasite Plasmodium falciparum. Here, we present functional and crystallographic data on P. falciparum prolyl-tRNA synthetase (PfPRS). Using immunofluorescence data, we show that PfPRS is exclusively resident in the parasite cytoplasm within asexual blood stage parasites. The inhibitor HF interacts strongly with PfPRS in a non-competitive binding mode in presence or absence of ATP analog. Intriguingly, the two monomers that constitute dimeric PfPRS display significantly different conformations in their active site regions. The structural analyses presented here provide a framework for development of febrifugine derivatives that can seed development of new anti-malarials.     

Phenotypic and genotypic characteristics of Arcanobacterium haemolyticum isolated from clinical samples in a Danish hospital

Six Arcanobacterium haemolyticum strains isolated from six patients of two hospitals in Denmark were identified phenotypically, also including matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis, and by genotypic methods. The latter were performed by sequencing 16S rDNA and glyceraldehyde 3-phosphate dehydrogenase encoding gene gap and by amplification of an A. haemolyticum specific region of 16S–23S rDNA intergenic spacer region and 23S rDNA. The six A. haemolyticum strains were further investigated for the presence of seven potential virulence genes encoding arcanolysin, phospholipase D, hemolysin A, CAMP factor family protein, collagen binding protein, neuraminidase A and neuraminidase H which appeared to be present in two (seven virulence genes), two (six virulence genes) and two strains (four virulence genes), respectively. The phenotypic and genotypic properties described in the present study might help to reliably identify and further characterize A. haemolyticum isolated from human patients, a species which seems to be of increasing importance.     

Identification of nucleolus-localized PTEN and its function in regulating ribosome biogenesis

The tumor suppressor PTEN is a lipid phosphatase that is found mutated in different types of human cancers. PTEN suppresses cell proliferation by inhibiting the PI3K-Akt signaling pathway at the cell membrane. However, PTEN is also demonstrated to localize in the cell nucleus where it exhibits tumor suppressive activity via a different, unknown mechanism. In this study we report that PTEN also localizes to the nucleolus and that nucleolar PTEN plays an important role in regulating nucleolar homeostasis and maintaining nucleolar morphology. Overexpression of nuclear PTEN in PTEN null cells inhibits Akt phosphorylation and reduces cell size. Knockdown of PTEN in PTEN positive cells leads to nucleolar morphologic changes and an increase in the proportion of cells with a greater number of nucleoli. In addition, knockdown of PTEN in PTEN positive cells increased ribosome biogenesis. These findings expand current understanding of function and relevance of nuclear localized PTEN and provide a foundation for the development of novel therapies targeting PTEN.     

Monoclonal antibody reveals H-2-linked quantitative and qualitative variation in the expression of a Qa-2 region determinant

We have produced a monoclonal antibody, Y-7, that reacts with a Qa-2 region-controlled determinant. Cellular and strain distribution analyses, coupled with quantitative variation in the amount of Y-7 antigen expressed among strains, provide overwhelming evidence that Y-7 reacts with the Qa-2^a determinant. The determinant detected by Y-7 is differentially expressed in T and B lymphocytes in a strain specific manner. Y-7 reacts with the majority of T lymphocytes (> 95%) and approximately one-half of B lymphocytes in certain strains (++ strains), and with the majority of T lymphocytes (> 95%) and no B lymphocytes in other strains (+ strains). T lymphocytes in + strains express approximately three fold less of the Y-7 determinant than T lymphocytes from ++ strains. In addition, we show that the Y-7 determinant is expressed in approximately one-third to one-half of Lyb-3^?, 5^? B lymphocytes. Possible mechanisms determining quantitative and qualitative variation in the expression of the Y-7 determinant in T and B lymphocytes are discussed.     

Cleft palate susceptibility linked to histocompatibility-2 (H-2) in the mouse

Data have been obtained indicating that cortisone-induced cleft palate in the mouse is linked to theH-2 ^a complex. Cortisone (2.5 mg) was administered to pregnant females on days 11 through 14 of pregnancy. On day 17 of pregnancy, the fetuses were inspected for cleft palates. Sham experiments were done by injecting sterile saline instead of cortisone. The inbred strains, A/J and C57BL/6, and the congenic strains C57BL/10ScSn and B10.A were tested for susceptibility to cleft palate. The clefting frequency was also observed in hybrids of the congenic strains. The A/J and B10.A strains showed a characteristic high susceptibility to cleft palate (i.e., 99% and 81% incidence of cleft palate, respectively) after teratogenic treatment. The C57BL/6 and C57BL/ 10ScSn demonstrated a significant resistance to the teratogen (i.e., 25% and 21 % incidence of clefting, respectively). The teratogenic treatment of congenic hybrids indicated that maternal influences significantly affected the incidence of cleft palate formation. The maternal influence appeared to depend upon the specificH-2 haplotype of the mother.     

Protoplast fusion enhances lignocellulolytic enzyme activities in Trichoderma reesei

Protoplast fusion was used to obtain a higher production of lignocellulolytic enzymes with protoplast fusion in Trichoderma reesei. The fusant strain T. reesei JL6 was obtained from protoplast fusion from T. reesei strains QM9414, MCG77, and Rut C-30. Filter paper activity of T. reesei JL6 increased by 18 % compared with that of Rut C-30. β-Glucosidase, hemicellulase and pectinase activities of T. reesei JL6 were also higher. The former activity was 0.39 Uml^?1, while those of QM9414, MCG77, and Rut C-30 were 0.13, 0.11, and 0.16 Uml^?1, respectively. Pectinase and hemicellulase activities of JL6 were 5.4 and 15.6 Uml^?1, respectively, which were slightly higher than those of the parents. The effects of corn stover and wheat bran carbon sources on the cellulase production and growth curve of T. reesei JL6 were also investigated.     

Pranceacanthus coccineus (Acanthaceae), a new genus and species from Amazonian brazil

The new genusPranceacanthus, apparently most closely related to the bitypic genusJuruasia, is described and illustrated. Its sole species,P. coccineus, has been collected on “terra firma” in lowland Amazonian Brazil in the states of Amazonas and Mato Grosso and in the territory of Rondônia.     

Glyoxylate conversion by Hyphomicrobium species grown on allantoin as nitrogen source

Glyoxylate, formed as a result of allantoin degradation, is converted by Hyphomicrobium species to glycerate via tartronate semialdehyde. Glyoxylate carboligase and tartronate semialdehyde reductase, the two enzymes involved, are present only in cells grown on allantoin as nitrogen source.     

Genetic basis for the polymorphism of rat liver cytosolic aldehyde dehydrogenase

Liver aldehyde dehydrogenase (ALDH), the enzyme involved in the oxidation of aldehydes such as acetaldehyde derived from ethanol, exists in multiple forms in most mammals. Up to five separable forms have been identified from the cytosolic fraction of Wistar rat liver. We investigated the genetic basis of a particular set of three enzyme forms by selective breeding and analysis of electrophoretic patterns of liver ALDH by isoelectric focusing. The forms of liver ALDH investigated were at pI 5.8 or 6.2, or a triple form with enzymes at pI 5.8, 6.0, and 6.2. There are two alleles found at the ALDH locus which encode in homozygotes for one of two electrophoretically separable ALDH forms. A rat heterozygous at the locus forms both ALDH types plus a hybrid. The alleles are expressed codominantly, found at an autosomal locus, and remain constant postpartum. The activities associated with the triplet enzyme form were statistically indistinguishable from a 1:2:1 ratio. This suggests that the enzymes hybridize to form a set of dimers or tetramers of the form A₂, AB, B₂ or A₄, A₂B₂, B₄, respectively.     

Influence of bovine lactoferrin on the growth of selected probiotic bacteria under aerobic conditions

Bovine lactoferrin (bLf) is a natural glycoprotein, and it shows broad-spectrum antimicrobial activity. However, reports on the influences of bLf on probiotic bacteria have been mixed. We examined the effects of apo-bLf (between 0.25 and 128 mg/mL) on both aerobic and anaerobic cultures of probiotics. We found that bLf had similar effects on the growth of probiotics under aerobic or anaerobic conditions, and that it actively and significantly (at concentrations of >0.25 mg/mL) retarded the growth rate of Bifidobacterium bifidum (ATCC 29521), B. longum (ATCC 15707), B. lactis (BCRC 17394), B. infantis (ATCC 15697), Lactobacillus reuteri (ATCC 23272), L. rhamnosus (ATCC 53103), and L. coryniformis (ATCC 25602) in a dose-dependent manner. Otherwise, minimal inhibitory concentrations (MICs) were 128 or >128 mg/mL against B. bifidum, B. longum, B. lactis, L. reuteri, and L. rhamnosus (ATCC 53103). With regard to MICs, bLf showed at least four-fold lower inhibitory effect on probiotics than on pathogens. Intriguingly, bLf (>0.25 mg/mL) significantly enhanced the growth of Rhamnosus (ATCC 7469) and L. acidophilus (BCRC 14065) by approximately 40–200 %, during their late periods of growth. Supernatants produced from aerobic but not anaerobic cultures of L. acidophilus reduced the growth of Escherichia coli by about 20 %. Thus, bLf displayed a dose-dependent inhibitory effect on the growth of most probiotic strains under either aerobic or anaerobic conditions. An antibacterial supernatant prepared from the aerobic cultures may have significant practical use.     

Study of enhancement and inhibition phenomena and genes relating to degradation of petroleum polycyclic aromatic hydrocarbons in isolated bacteria

Polycyclic aromatic hydrocarbons (PAHs) are xenobiotic compounds, which being degraded by chemical, physical or biological methods. The latter is the safest and the cheapest one. Two bacterial strains ASU-01 and ASU-016 were isolated from different Egyptian petroleum contaminated sites. They were genetically identified based on the analysis of the nucleotide sequences of the 16S ribosomal PNA gene and the phylogenetic tree as Enterobacter hormaechei and Pseudomonas pseudoalcaligenes respectively. When pyrene as high molecular weight (HMW)-PAH was added as a sole carbon source, both strains could degrade it with efficiency 77.7 and 83.7% within 15 days of incubation, respectively. However, when it was mixed with low molecular weight (LMW)-PAHs, two opposite phenomena appeared. The first one was enhancement, which occurred with ASU-01. This strain shifted pyrene efficiency to 98.5%. The second phenomenon was inhibition occurred with ASU-016 which completely retarded pyrene degradation. Naphthalene dioxygenase (nahAc), and catechol dioxygenases (C12O and C23O) genes were detected in the two strains based on PCR. The detected genes were confirmed by determining the different specific activities of their translated protein (enzymes) on different PAHs. The maximum values of biosurfacatant production activity and cell-surface and percentage of cell-surface hydrophobicity (CSH) were detected during the exponential phase. These latter factors increased the bioavailability and consequently, the assimilation of PAHs.