Distribution of chitinase and chitobiase in bacillus

Sixty strains representing 29 taxospecies ofBacillus were assayed for their ability to hydrolyze colloidal chitin. A qualitative estimation of chitinolysis was made from the clear zone produced around colonies in the conventional agar plate method and chitobiase activity by use of the fluorescence of 4-methylumbelliferyl-N-acetyl--d-glucosaminide.Strains positive in the chitin-agar plate method were assayed for production of reducing sugar in liquid culture. Seventeen of 52 strains representing 10 species ofBacillus were chitinolytic. The most chitinolytic species ofBacillus were:B. chitinosporus, B. pulvifaciens, B. alvei, B. Macerans, andB. licheniformis. Seventy-eight percent ofBacillus isolates from chitinenriched soil (AU Y91B1, AU-X (unidentified), and AU B₂–B₈) were chitinolytic. Twenty-three strains representing 15 species gave a positive test for chitobiase. Many strains negative for endochitinase gave a strong positive reaction (4+) for chitobiase.     

Microsatellite markers isolated from Drosophila hydei

We isolated and characterized 10 polymorphic microsatellite loci in Drosophila hydei. The number of alleles per locus ranged from 3 to 8 (N = 23 individuals). Polymorphic information content ranged from 0.316 to 0.750 and observed heterozygosity from 0.261 to 0.913. These markers will be valuable in studies of sexual selection and parental investment in D. hydei.     

Characterization of cloned ribosomal DNA from Drosophila hydei.

The structure of ribosomal genes from the fly Drosophila hydei has been analyzed. EcoRI fragments, cloned in a plasmid vector, were mapped by restriction enzyme analysis. The lengths of the regions coding for 18S and 28S rRNA were defined by R-loop formation. From these data a physical map of the rRNA genes was constructed. There are two major types of rDNA units in D. hydei, one having a size of 11 kb and the other a size of 17 kb. The 17 kb unit results from an intervening sequence (ivs) of 6.0 kb, interrupting the beta-28S rRNA coding region. Some homology between th D. hydei ivs and D. melanogaster type 1 ivs has been described previously (1). However, the restriction sites within these ivs show considerable divergence. Whereas D. hydei rDNA D. melanogaster rDNA, the nontranscribed spacer has little, if any, sequence homology. Despite difference in sequence, D. hydei and D. melanogaster spacers show structural similarities in that both contain repeated sequence elements of similar size and location.     

Purification and molecular characterization of alcohol dehydrogenase from Drosophila hydei: Conservation in the biochemical features of the enzyme in several species of Drosophila

Alcohol dehydrogenase (ADH) has been purified from Drosophila hydei. Biochemical investigations show that the native enzyme is a dimer consisting of two identical subunits with molecular weight 27,000. The pH optimum values of pure enzyme preparations are 7.9 and 9.4. The pI values are 8.83 and 8.41. Substrate specificities have been characterized. Km(app) values are lowest for propan-2-ol and butan-2-ol and Vmax(app) values are highest for these two substrates. The amino acid composition has been determined. Peptide mapping experiments performed after trypsin digestion of the enzyme allow the identification of 24 peptides. Peptides comprising 64% of the amino acid residues have also been purified by high-performance liquid chromatography (HPLC), and their N-terminal residues and amino acid composition determined. Results are compared with the amino acid sequence of ADH from D. melanogaster Adhs [Thatcher, D. R. (1980). Biochem. J. 187:875]. When data on the biochemical and structural characterization of ADH from D. hydei are compared with data from other species of Drosophila, clear homologies are observed.     

Y chromosomal DNA of Drosophila hydei

Six recombinant DNA clones are described, which are derived from the Y chromosome of Drosophila hydei. They reveal characteristic features of Y chromosomal DNA sequences. Three of the cloned inserts are Y-specific and are members of the same family of repeated sequences associated with the lampbrush loop-forming fertility gene "nooses" in the short arm of the Y chromosome. The other three cloned sequences are members of three different families of repeated sequences, but display a small amount of homology to one another and to the family of the nooses sequences. These three cloned sequences are found preferentially in the Y chromosome, but also in other chromosomal positions. The Y chromosomal copies are located in the short arm of the Y chromosome. The other copies are found in autosomal kinetochore-associated heterochromatin or, for one of the cloned sequences, in one band of the giant chromosome 4, in addition to the kinetochore heterochromatin.     

The ribosomal DNA of Drosophila melanogaster is organized differently from that of Drosophila hydei

On the X chromosome of Drosophila melanogaster there is a single tandem array of 240 ribosomal RNA genes. The majority of these contain an insertion, known as type I, in the 28 S coding region. Previous genetic and electron microscopic studies indicated that genes bearing the type I insertion (ins⁺) are interspersed at random with those lacking it (ins^?). In contrast, Renkawitz-Pohl et al. (1981) have analyzed the restriction pattern of X chromosomal ribosomal DNA in Drosophila hydei and demonstrated that in this case ins⁺ genes are segregated from ins^?. This suggests either that the rDNA is organized differently in these two species or that the restriction enzyme technique reveals significant clustering not detected by previous methods. By using an appropriate restriction enzyme, we demonstrate that ins⁺ and ins^? genes are intermingled at random in D. melanogaster. These experiments also indicate that genes containing the short form of the insertion are flanked by a larger spacer upstream than downstream.     

RNA metabolism during metamorphosis of Drosophila hydei

Determinations of total protein and RNA at various times after the beginning of the pharate pupal stage of Drosophila hydei, revealed an increase in both substances during the first 25 hr and a sudden decrease thereafter until 52 hr. From this time on the total amount of both protein and RNA increases slightly until emergence of the flies at 160 hr after the beginning of the pharate pupal stage. A similar pattern of changes was recorded for the total radioactivity as well as the specific activity of RNA labelled with ³H-uridine after the injection of the isotope immediately before the beginning of the pharate pupal stage.The migration profile of RNA labelled with ³H-uridine during larval development, revealed that shortly after the onset of the pharate pupal stage an essentially normal larval pattern consisting of major fractions of 28, 18, 8 to 9, and 4 to 7 S RNA. At 52 hr only the low molecular weight RNA was present. The ‘normal’ pattern was restored at the time of emergence of the flies, indicating re-utilization of degradation products of previously labelled RNA.     

Purification and characterization of an extracellular chitobiase fromTrichoderma harzianum

Chitobiase (EC 3.2.1.29), from the culture filtrate ofTrichoderma harzianum, was purified in sequential steps by ammonium sulfate precipitation, ion exchange chromatography, and gel filtration. The physical and biochemical properties of the enzyme have been determined. The native enzyme has a molecular weight of 118 kDa when determined by gel filtration, and 64 kDa by SDS-PAGE. The enzyme catalyzed the hydrolysis of N,N-diacetylchitobiose andp-nitrophenyl--N-acetyl glucosamine with apparent Km of 575 µM and 235 µM, respectively. The pH optimum for the enzyme was pH 5.5, and maximum activity was obtained at 50°C. Glucosamine and N-acetylglyucosamine strongly inhibited the enzyme.     

Purification and characterization of chitinase from Alcaligenes xylosoxydans

Extracellular chitinase from Alcaligenes xylosoxydans was purified to electrophoretic homogeneity using affinity and gel filtration chromatography. The molecularmass of chitinase was estimated to be 45 kDa and44 kDa by SDS-PAGE and gel-filtration, respectively. The enzyme was optimally active at 50 °C (over 30 min) and pH 5. Activity staining after PAGE showed a single band. The Km for chitin was 3 g l^–1. Cu²⁺ and Na⁺ at 5 mM inhibited chitinase activity to 25% while Ca²⁺, Mg²⁺ and Ba²⁺ had no effect at the same concentration. The purified enzyme degraded mycelia of Aspergillus niger.     

The action of the Notch locus in Drosophila hydei

The phenotypic effects of different doses of the dominant, sex-linked mutant Notch (N) and its wildtype allele (N ⁺) were studied in Drosophila hydei, N being lethal in homozygous or hemizygous condition. Various dosage combinations were made by using N ⁺ N and N ⁺ N ⁺ attached-X chromosomes as well as X and Y N ⁺-duplication chromosomes (w ^mCoY, Xw^mCo,and DpCo ^Nt). The N mutant used, N ⁶⁸, is associated with a small inversion: In (I) N ⁶⁸.The wing phenotype was found to depend solely on the number of functional (N ⁺) alleles present, irrespective of the dose of N. Females with a single dose of N ⁺ are phenotypically Notch, females with three or four doses of N ⁺ show a Confluens wing phenotype. The latter occurs in varying degrees of expression which seem to be correlated with the relative amounts of sex-chromosomal heterochromatin present. In males the N ⁺ locus behaves as a dosage compensated locus either on the X or the Y chromosome.In the w ^mCo (w⁺N⁺) duplication, the w ⁺ locus shows variegation when placed over white, whereas N ⁺ placed over N ⁶⁸ does not. The former being situated closer to the heterochromatin in this aberration, this is consistent with the idea of gene inhibition by heterochromatin but at the same time would imply a very limited spreading effect.