Characterization of Melanocarpus albomyces steryl esterase produced in Trichoderma reesei and modification of fibre products with the enzyme

Melanocarpus albomyces steryl esterase STE1 is considered to be an interesting tool for several industrial applications due to its broad substrate specificity. STE1 was produced in the filamentous fungus Trichoderma reesei in a laboratory bioreactor at an estimated production level of 280 mg l^−l. The properties of the purified recombinant enzyme (rSTE1), such as substrate specificity, molecular mass, pH optimum and stability and thermostability, were characterized and compared to the corresponding properties of the native enzyme. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed one band with a molecular weight of 60 kDa for rSTE1, whereas analytical gel filtration showed a dimeric structure with a molecular weight of 120 kDa. The rSTE1 was somewhat less stable under different conditions and had slightly lower activities on various substrates than the native STE1. The effects of rSTE1 on the properties of paper sheets and polyethylene terephthalate (PET) fabric were preliminarily evaluated. Due to the hydrolysis of triglycerides and steryl esters by the rSTE1 treatment, the tensile strength and hydrophilicity of the paper were increased. The rSTE1 treatment increased significantly the polarity of PET by hydrolysing the ester bonds in the polyester backbone. Dyeing of PET with methylene blue was also slightly improved after rSTE1 treatment.     

Egg Carrying Attracts Enemies in a Cryptic Coreid Bug (Phyllomorpha laciniata )

In the golden egg bug (Phyllomorpha laciniata) eggs are laid mainly on the backs of conspecifics, and in many habitats eggs do not survive unless carried by bugs. Bugs are covered with small spines that may make them unpalatable. They are also cryptic, at least if not carrying eggs. We used domestic chicks as predators to examine if egg carrying influences susceptibility to avian predators. The special morphology of the bugs and/or possible chemical defense may make the bugs unattractive, as chicks that picked up bugs often rejected them. Eggs made bugs more attractive to chicks. The total number of attacks and the probability of being attacked at all increased significantly when bugs carried eggs. If mating with an egg-carrying bug, a female without eggs suffered as much as her egg-carrying partner when attacked. This study, together with previous results on ant predation, suggests that carrying eggs as well as mating with an egg-loadedbug are costly in terms of predation risk.     

Protein-bound oligosaccharides of Semliki Forest virus

Semliki Forest virus was grown in BHK cells and labeled in vivo with radio-active monosaccharides. promnase digenst of the virus chromatographer on Bio-Gel P 6 revealed glycopeptides of A-type and B-type. (For the nomenclature see Johnson J. and Clamp J.R. (1971) Biochem. J. 123, 739–745) The former was labeled with [³H]fucose, [³H]galactose, [³H]mannose and [¹⁴C]glucosamine, the latter only with [³H]mannose and [¹⁴C]glucosamine. The three envelope glycoproteins E₁, E₂ and E₃ were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subjected to pronase digestion. The glycoproteins E₁ and E₃ revealed glycopeptides of A-type. E₂ revealed glycopeptides of B-type. E₂ yielded additionally a glycopeptide (M_r3100) which was heavily labeled from [³H]galactose, but only marginally from [¹⁴C]glucosamine, [³H]fucose and [³H]mannose. Wether this glycopeptide belongs to the A-type or not remains uncertain. The apparent molecular weights of the A-type units measured by gel filtration were 3400 in E₁ and 4000 in E₃; the B-type unit of E₂ had an apparent molecular weight of 2000. Combined with the findings of our earlier chemical analysis these data suggast that E₁ and E₃ contain on the average one A-type unit; E₂ probably contains one 3100 dalton unit plus one or two B-type units.     

High frequency one-step gene replacement in Trichoderma reesei. II. Effects of deletions of individual cellulase genes

Four cellulase genes of Trichoderma reesei, cbh1, cbh2, egl1 and egl2, have been replaced by the amdS marker gene. When linear DNA fragments and flanking regions of the corresponding cellulase locus of more than 1 kb were used, the replacement frequencies were high, ranging from 32 to 52%. Deletion of the major cellobiohydrolase 1 gene led to a 2-fold increase in the production of cellobiohydrolase II; however, replacement of the cbh2 gene did not affect the final cellulase levels and deletion of egl1 or egl2, slightly increased production of both cellobiohydrolases. Based on our results, endoglucanase II accounts for most of the endoglucanase activity produced by the hypercellulolytic host strain. Furthermore, loss of the egl2, gene causes a significant drop in the filter paper-hydrolysing activity, indicating that endoglucanase II has an important role in the total hydrolysis of cellulose.     

Genome-wide signatures of differential DNA methylation in pediatric acute lymphoblastic leukemia

Background

Although aberrant DNA methylation has been observed previously in acute lymphoblastic leukemia (ALL), the patterns of differential methylation have not been comprehensively determined in all subtypes of ALL on a genome-wide scale. The relationship between DNA methylation, cytogenetic background, drug resistance and relapse in ALL is poorly understood.

Results

We surveyed the DNA methylation levels of 435,941 CpG sites in samples from 764 children at diagnosis of ALL and from 27 children at relapse. This survey uncovered four characteristic methylation signatures. First, compared with control blood cells, the methylomes of ALL cells shared 9,406 predominantly hypermethylated CpG sites, independent of cytogenetic background. Second, each cytogenetic subtype of ALL displayed a unique set of hyper- and hypomethylated CpG sites. The CpG sites that constituted these two signatures differed in their functional genomic enrichment to regions with marks of active or repressed chromatin. Third, we identified subtype-specific differential methylation in promoter and enhancer regions that were strongly correlated with gene expression. Fourth, a set of 6,612 CpG sites was predominantly hypermethylated in ALL cells at relapse, compared with matched samples at diagnosis. Analysis of relapse-free survival identified CpG sites with subtype-specific differential methylation that divided the patients into different risk groups, depending on their methylation status.

Conclusions

Our results suggest an important biological role for DNA methylation in the differences between ALL subtypes and in their clinical outcome after treatment.     

Production in <Emphasis Type="Italic">Trichoderma reesei</Emphasis> of three xylanases from <Emphasis Type="Italic">Chaetomium thermophilum</Emphasis>: a recombinant thermoxylanase for biobleaching of kraft pulp

Three endoxylanase genes were cloned from the thermophilic fungus Chaetomium thermophilum CBS 730.95. All genes contained the typical consensus sequence of family 11 glycoside hydrolases. Genomic copies of Ct xyn11A, Ct xyn11B, and Ct xyn11C were expressed in the filamentous fungus T. reesei under the control of the strong T. reesei cel7A (cellobiohydrolase 1, cbh1) promoter. The molecular masses of the Ct Xyn11A, Ct Xyn11B, and Ct Xyn11C proteins on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were 27, 23, and 22 kDa, respectively. Ct Xyn11A was produced almost as efficiently as the homologous xylanase II from a corresponding single-copy transformant strain. Ct Xyn11B production level was approximately half of that of Ct Xyn11A. The amount of Ct Xyn11C was remarkably lower. Ct Xyn11A had the highest temperature optimum and stability of the recombinant xylanases and the highest activity at acid-neutral pH (pH 5–7). It was the most suitable for industrial bleaching of kraft pulp at high temperature.     

Females show greater changes in wing colour with latitude than males in the green‐veined white butterfly,Pieris napi (Lepidoptera: Pieridae)

In butterflies, wing colour may simultaneously be under sexual selection in the context of mating selection and natural selection in the context of thermoregulation. In the present study, we collected mated females of the green‐veined white butterfly (Pieris napi) from locations spanning 960 km of latitude across Fennoscandia, and investigated sex‐specific latitudinal wing colour variation in their offspring raised under identical conditions. We measured wing colour characteristics, including reflectance at wavelengths 300–700 nm and the degree of wing melanization. At all latitudes, females reflected more light in the short wavelengths (< 400 nm) and less in the long wavelengths (> 450 nm), and they were more melanized than males. However, female wing colour varied more with latitude than that of males. Among females, long wavelength reflectance decreased, whereas short wavelength reflectance and melanization increased, towards the north. By contrast, among males, latitudinal variation was found only in the ventral hindwing melanization. These results are consistent with the idea that the balance between natural and sexual selection acting on wing colour changes with latitude differently in males than females. The dark wing colour of females in the north may be a thermoregulatory adaptation, although males may be constrained from evolving the dark dorsal wing colour favoured by natural selection because of constant sexual selection across latitudes. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.     

Comparison between the preservation of chloroplast structure in the dark and the turnover rate of chlorophyll-protein complexes in a moss and two varieties of pea

The senescence and preservation of the structure of the chloroplastin the dark was studied in the protonema of Ceratodon purpureusand in two varieties of pea, a tall variety "Dippe Maj" anda dwarf variety "Early low". The latter variety is better ableto survive in darkness, probably because of the larger amountof starch in its chloroplasts. The integrity of the chloroplastswas better preserved in the moss than in the peas. During isolationof the chlorophyll-protein complexes, there was no decreasein the specific activity of the total lamellar extract in anyof the dark-grown materials. This contrasts with the turnoverof chlorophyll-protein complex (CP) II in the dark-grown moss.In the light, CP II had a half life of two weeks in both themoss and the higher plants, whereas no turnover was evidentin CP I. The lack of turnover for CP I is connected with itsdecrease in senescence. ¹Present address: Department of Botany, University of Turku,20500 Turku 50, Finland. (Received July 13, 1979; )     

Soluble and immobilized enzyme technology in bioconversion of barley starch

Homogeneous and heterogeneous biocatalysis were both investigated as tools for barley starch syrup production. Barley starch was first liquefied by soluble heat-stable Bacillus sp. α-amylase EC 3.2.1.1 (1,4-α-d-glucan glucanohydrolase) Termamyl 60 L at 95°C, pH 6.5, to obtain slurries of varying DE-values up to ≈37. Alternatively, it was extruded with a Creusot-Loire BC 45 twin-screw extruder at 25% moisture, 150°C, for denaturation. After cooling and adjusting the pH to 4.5 or grinding, respectively, the pretreated starch was saccharified either by soluble or by immobilized Aspergillus niger glucoamylase EC 3.2.1.3 (1,4-α-d-glucan glucohydrolase) at 60°C, pH 4.5, to obtain glucose syrup of up to DE 96. The course of hydrolysis was followed by automated Biogel P-2 chromatographic analysis. Glucoamylase was immobilized either on a phenol-formaldehyde resin Duolite S 761 or on silanized Spherosil porous silica beads. Barley glucose syrup obtained was further continuously converted to high fructose syrup by a packed bed reactor of Actinoplanes missouriensis whole cell glucose isomerase (EC 5.3.1.5) Maxazyme entrapped within α-cellulose beads. We could conclude that barley starch may be used as an alternative raw material for biocatalytic starch syrup production.