Identification of a second DNA binding site in the human Rad52 protein

Rad52 plays essential roles in homology-dependent double-strand break repair. Various studies have established the functions of Rad52 in Rad51-dependent and Rad51-independent repair processes. However, the precise molecular mechanisms of Rad52 in these processes remain unknown. In the present study we have identified a novel DNA binding site within Rad52 by a structure-based alanine scan mutagenesis. This site is closely aligned with the putative single-stranded DNA binding site determined previously. Mutations in this site impaired the ability of the Rad52-single-stranded DNA complex to form a ternary complex with double-stranded DNA and subsequently catalyze the formation of D-loops. We found that Rad52 introduces positive supercoils into double-stranded DNA and that the second DNA binding site is essential for this activity. Our findings suggest that Rad52 aligns two recombining DNA molecules within the first and second DNA binding sites to stimulate the homology search and strand invasion processes.     

Stereoselective synthesis of (22R)- and (22S)-castasterone/ponasterone A hybrid compounds and evaluation of their molting hormone activity

Two stereoisomers of a castasterone/ponasterone A hybrid compound, the (20R,22R) and (20R,22S)-isomers of 2alpha,3alpha,20,22-tetrahydroxy-5alpha-cholestan-6-one, were synthesized stereoselectively and their binding activity to the ecdysteroid receptor was determined. From the concentration-response curve for the inhibition of the incorporation of tritiated ponasterone A into ecdysteroid receptor containing insect cells, the concentration (IC50) required to inhibit 50% of the incorporation of radioactivity into cells was evaluated. The IC50 values of the (22R)- and (22S)-isomers were determined to be 0.30 and 38.9 microM against Kc cells, respectively, indicating that the (22R)-isomer is about 100 times more potent than the corresponding (22S)-isomer. IC50 values of these compounds against lepidopteran Sf-9 cells were determined to be 0.36 and 12.9 microM, respectively. The molting hormonal effect was examined in a Chilo suppressalis integument system and the 50% effective concentration for the stimulation of N-acetylglucosamine incorporation into the cultured integument was determined to be 2.7 microM for the (22R)-isomer, while the (22S)-isomer was inactive. On the other hand, both isomers did not show brassinolide-like activity in the rice lamina inclination assay.     

Cleome hassleriana plants fully support the development and reproduction of Nesidiocoris tenuis

BioControl - Nesidiocoris tenuis (Reuter) (Hemiptera: Miridae) is a zoophytophagous predator that feeds on plants as well as prey. Several non-crop host plant species have been used to maintain...     

The Chromatographic Purification of Yeast Invertase by an Ion-Exchange Resin Method and Some Properties of the Enzyme Obtained

The purification of yeast invertase was attempted by application of the chromatographic method using Duolite C-10, a sulfonic acid cation exchange resin. This method was found to be extremely simple in process and significantly effective for the improvement of purity of the enzyme, compared with those other methods reported, hitherto. In the present paper, the procedure of the purification and some properties of the enzyme obtained thereby, are described, and some discussion of the implications is presented.     

Studies on Urate Oxidase of Candida utilis

Some physical and chemical properties of urate oxidase (EC 1.7.3.3) isolated from the cells of Candida utilis were investigated. The molecular weight was estimated to be 1.2 × 10⁵ by the equilibrium sedimentation and gel filtration methods. The isoelectric point was determined as 5.4 by the method of density electrofocusing. The enzyme showed a slight absorption at 410 mμ, and the absorbancy at this wave length was only 3% of that at 280 mμ. Contrary to urate oxidase from swine liver, the enzyme from yeast contained a negligible amount of copper, but it contained iron of nearly one atom per mole of the enzyme protein. The yeast urate oxidase was not inactivated by some chelators. However, it was easily inactivated with certain heavy metal ions such as Hg²⁺, and the inactivated enzyme was reactivated by the addition of thiols, indicating that the enzyme is a sulfhydryl enzyme. The inactivation of the enzyme with urea, on the other hand, was greatly accelerated by the addition of thiols, and some discussion was added to the results obtained.     

α-Amylase Formation and Nucleic Acid Metabolism of Bacillus subtilis

The nucleic acid metabolism in washed cells of Bacillus subtilis was investigated with special reference to amylase formation of the bacterium. On incubation of the suspension of the washed cells, purines, pyrimidines and their related compounds were observed in the medium. However, in the medium of the cells incubated with a calcium chelater, where no amylase formation occurred, were detected adenosine- and guanosine-monophosphate in addition to those described above. The addition of a calcium chelater was also found to decrease the quantity of the nucleic acids being involved in the lysozyme-sensitive fraction of the bacterial cells, suggesting the possibility that the metabolism of nucleic acids in this fraction is closely related to amylase formation of the cells.     

Validation of treatments for conservation of native rhododendrons, the symbol of Satoyama, in the dry granite region of Japan

To develop an appropriate method of conservation for native Rhododendron sections Brachycalyx and Tsutsusi, the symbol of Satoyama, field experiments were performed in the dry granite region of Japan. When carpet-type landscape of native rhododendrons was desired, all plants were cut at 20 cm above the ground level of all trees and shrubs, and herbs and ferns were weeded (once and three times, respectively). When shrubby-type landscape of native rhododendrons was desired, all plants, excluding native rhododendrons, were cut at 20 cm above the ground level of all trees and shrubs, herbs and ferns were weeded, and litter was swept. After 3 years of monitoring of the percentage and depth of crowns with flower buds, the following major results were obtained: Cutting all plants excluding native rhododendrons was effective to maintain the depth of crowns with flower buds. However, weeding and sweeping of litter on the ground caused desiccation of surface soil, which induced a transitory decrease in the percentage of crowns with flower buds. One weeding was effective in maintaining the depth of crowns with flower buds; however, the second and third weedings had no distinct effect. In dry granite regions, considerable attention to desiccation of surface soil is critical, as opposed to limiting attention to maintenance of sunlight as is common practice.     

Monitoring the rate and dynamics of concerted evolution in the ribosomal DNA repeats of Saccharomyces cerevisiae using experimental evolution

Concerted evolution describes the unusual evolutionary pattern exhibited by certain repetitive sequences, whereby all the repeats are maintained in the genome with very similar sequences but differ between related species. The pattern of concerted evolution is thought to result from continual turnover of repeats by recombination, a process known as homogenization. Approaches to studying concerted evolution have largely been observational because of difficulties investigating repeat evolution in an experimental setting with large arrays of identical repeats. Here, we establish an experimental evolution approach to look at the rate and dynamics of concerted evolution in the ribosomal DNA (rDNA) repeats. A small targeted mutation was made in the spacer of a single rDNA unit in Saccharomyces cerevisiae so we could monitor the fate of this unit without the need for a selectable marker. The rate of loss of this single unit was determined, and the frequency of duplication was also estimated. The results show that duplication and deletion events occur at similar rates and are very common: An rDNA unit may be gained or lost as frequently as once every cell division. Investigation of the spatial dynamics of rDNA turnover showed that when the tagged repeat unit was duplicated, the copy predominantly, but not exclusively, ended up near to the tagged repeat. This suggests that variants in the rDNA spread in a semiclustered fashion. Surprisingly, large deletions that remove a significant fraction of total rDNA repeats were frequently found. We propose these large deletions are a driving force of concerted evolution, acting to increase homogenization efficiency over-and-above that afforded by turnover of individual rDNA units. Thus, the results presented here enhance our understanding of concerted evolution by offering insights into both the spatial and temporal dynamics of the homogenization process and suggest an important new aspect in our understanding of concerted evolution.