The stability and aggregation properties of human liver acid beta-D-galactosidase.

1. A method is described for following continuously the action of beta-galactosidase on 4-methylumbelliferyl beta-D-galactoside at pH 4.5, in which 4-methylumbelliferone production is measured at fluorescence excitation and emission wavelengths of 324 and 444nm respectively. 2. Initial-rate studies show that the presence of salt activates beta-galactosidase up to 100 mM, but is inhibitory above that concentration. The enzyme is very unstable at 37 degrees C and low ionic strength, but stability increases with ionic strength. 3. The stability of the enzyme at 37 degrees C decreases markedly with rising pH in the range 5.9--8.0. 4. Gel-filtration patterns demonstrate that there is a marked tendency to polymerization with increasing ionic strength. The gel-filtration pattern shows decreasing amounts of dimer with increasing pH. 5. The correlation between activity, stability and molecular form of beta-galactosidase is discussed. It is suggested that the dimeric form of the enzyme is the most stable and active form. The implications of this finding for the assay of beta-galactosidase under physiological conditions for prenatal diagnosis are discussed. 6. Evidence for the possible occurrence of a 36 000-mol.wt. from of beta-galactosidase is presented. 7. A computer program for the calculation of initial rates has been deposited as Supplementary Publication SUP 50114 (4 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1981) 193, 5.     

Embryogenesis of photoautotrophic cell cultures of Daucus carota L.

In this paper photoautotrophic carrot (Daucus carota L.) suspension cultures are described which are able to produce somatic embryos. The development of somatic embryos, however, requires a sucrose supplement. Although an elevation of the CO₂ concentration up to 2.3% results in the same level of dry weight production as with sucrose in the medium, somatic embryos could not be observed.Results on the influence of sucrose on some aspects of the photosynthetic apparatus of cultured cells are discussed.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - DW dry weight - ELISA enzyme-linked-immunosorbent-assay - FW fresh weight - IAA indole-3-acetic acid - NAA naphthaleneacetic acid - PEPCase phosphoenol-pyruvate carboxylase - Rubisco Ribulose- 1,5-bisphosphate carboxylase/oxygenase - se somatic embryogenesis     

Assimilatory Sulfate Reduction by the Hemoflagellate Leishmania tarentolae1

SYNOPSIS. Continuous growth of one cell line (UCI variant) of Leishmania tarentolae was achieved in the absence of organic sulfur. These cells were able to use sodium sulfate, and, to a limited extent, sodium sulfite as their sole sulfur source and could utilize methionine sulfoxide in place of L-methionine. A related cell line (RU variant) was unable to grow in organic sulfate-free media nor could these cells utilize methionine sulfoxide. UCI promastigotes incorporated significant amounts of ³⁵S sodium sulfate; killed cells did not take up the label. ³⁵S incorporation was inhibited by sodium molybdate (5 × 10^?4 M), sodium arsenite (5 × 10^?4 M), 2,4-dinitrophenol (1 × 10^?4 M), or KCN (5 × 10^?4 M). RU promastigotes did not incorporate significant amounts of ³⁵S sodium sulfate. Thin layer chromatographs of protein hydrolysates from UCI cells incubated in ³⁵S sodium sulfate revealed several radio opaque spots, one of which had chromatographic properties of cystine. UCI variants of L. tarentolae were therefore capable of assimilatory sulfate reduction whereas RU cells lacked this ability.     

Effects of nalidixic acid, chloramphenicol, cycloheximide, and anisomycin on structure and development of plastids and mitochondria in greening Euglena gracilis

The substructure of plastids and mitochondria and the alterations caused by the addition of antibiotics were investigated during light-induced proplastid-to-chloroplast transformation in Euglena gracilis. The organisms were grown in presence of the inhibitors up to 3 days (5 generations). Both 40 μg/ml nalidixic acid and 1–1.5 mg/ml chloramphenicol prevent the formation of chloroplasts of normal size and structure by blocking development during early stages. Under our conditions 2 to 5 straight thylakoids are formed beside 1 to 2 girdle-like thylakoids. The former rarely fuse into bands. Non-crystalline prolamellar bodies of considerable size are formed at the distended ends of the plastids in the presence of both drugs. Chloramphenicol also influences mitochondrial size, shape and internal structure. Giant mitochondria can be observed. Nalidixic acid does not change the size and shape of mitochondria, but the matrix frequently appears highly osmiophilic. Cycloheximide in sublethal doses (2–5 μg/ml) or 50 μg/ml anisomycin inhibits plastid development only in the early period after addition. In later culture periods chloroplasts are found enlarged in size with an increased number of thylakoids and bands per organelle. Insertions of new bands are noted at the inner membrane of the chloroplast envelope. The electronmicroscopic observations agree with the results of chloroplast-specific biochemical activities such as light-induced increase in chlorophyll synthesis and of two chloroplast-bound enzyme activities. The results are discussed with respect to metabolic and biogenetic correlations between the two types of organelles in E. gracilis cells.     

Graph-based clustering and characterization of repetitive sequences in next-generation sequencing data

Background  

The investigation of plant genome structure and evolution requires comprehensive characterization of repetitive sequences that make up the majority of higher plant nuclear DNA. Since genome-wide characterization of repetitive elements is complicated by their high abundance and diversity, novel approaches based on massively-parallel sequencing are being adapted to facilitate the analysis. It has recently been demonstrated that the low-pass genome sequencing provided by a single 454 sequencing reaction is sufficient to capture information about all major repeat families, thus providing the opportunity for efficient repeat investigation in a wide range of species. However, the development of appropriate data mining tools is required in order to fully utilize this sequencing data for repeat characterization.