A Zn2+-dependent and histone H1-specific protease in sea urchin sperm

Protease activity was extracted from sea urchin sperm with 1% Triton X-100 and partially purified by DEAE-cellulose and Sephadex G-100 chromatography. The enzyme preferentially degraded histone H1, while showing only a weak activity toward other histones. Heat-denatured casein and bovine serum albumin were not digested by this enzyme under the present experimental conditions. This protease hydrolyzed only Boc-Val-Leu-Lys-MCA among various peptidyl-MCAs. The optimal pH ranged from 7 to 11. Its molecular weight was about 41,000. Among various known inhibitors of proteases, only omicron-phenanthroline effectively inhibited the activity. The enzyme was stimulated by Zn2+ or Co2+. It was inactivated by omicron-phenanthroline but could be reactivated by the addition of Zn2+ or Co2+. Therefore, this protease seems to be a metalloprotease dependent on Zn2+ or Co2+. The insensitivity of this enzyme to phosphoramidon and its very restricted substrate specificity suggest that this enzyme is very different from other metalloproteases described hitherto.     

Mutation induction in Escherichia coli incubated in the reaction mixture of NADPH-dependent lipid peroxidation of rat-liver microsomes

Experiments were carried out to examine mutation induction in E. coli cells incubated in the reaction mixture of NADPH-dependent lipid peroxidation of microsomes isolated from rat liver. The results obtained were as follows: (1) Lipid peroxidation of microsomes occurred extensively on incubation with NADPH and Fe2+. In the E. coli WP2uvrA(pKM101) system, the mutation frequency to streptomycin resistance increased markedly when the cells were incubated in the reaction mixture of microsomal lipid peroxidation. The induced mutation frequencies were dependent on the extent of the lipid peroxidation. (2) It was also found that the mutations were induced at the same rate as in the case of (1) when the cells were added to the microsomal suspensions after the reactions due to the short-lived free radicals had terminated. (3) The cytotoxicity of the lipid peroxidation products was larger in the DNA repair-defective mutant, E. coli SR18 (uvrArecA) than the wild-type strain, SR749. From these results it is concluded that some DNA-damaging and mutagenic substances are indeed produced in the degradation process of peroxidized polyunsaturated fatty acids in liver microsomal lipids.     

Replication timing: histone genes replicate during early S phase in cleavage-stage embryos of sea urchin.

Newly synthesized DNA was separated from the bulk of the DNA by pulse-labeling with BUdR and centrifugation in an alkaline CsCl buoyant density gradient. The content of histone gene in the newly synthesized DNA was determined by DNA dot hybridization. The gene contents in DNA replicated during the early half of S phase and during the whole S phase were compared. Results showed that histone genes were replicated during the first half of the S phase in embryos in the early cleavage stage.     

Termini of Salmonella flagellin are disordered and become organized upon polymerization into flagellar filament

The terminal regions of Salmonella flagellin are essential for polymerization to form the flagellar filament. It has recently been suggested, on the basis of results from circular dichroism spectroscopy and scanning calorimetry, that these regions are disordered in solution. We report here direct evidence for disorder and mobility in the terminal regions of flagellin using 400 MHz proton nuclear magnetic resonance (n.m.r.) spectroscopy. Comparison of the n.m.r. spectra of monomeric and polymeric flagellin shows that the terminal regions become organized when polymerized to form the filament.     

Fusarium poae and Fusarium crookwellense, Fungi Responsible for the Natural Occurrence of Nivalenol in Hokkaido

To determine the reasons for the natural occurrence of nivalenol in the northernmost area of Japan, scabby wheat was harvested from 19 crop fields in Hokkaido. Mycological surveys and analysis for mycotoxin contamination were performed. Among 13 wheat grain samples harvested in seven locations, 9, 2, and 6 samples were positive for deoxynivalenol, nivalenol, and zearalenone, respectively, at levels ranging from 0.03 to 1.28 μg/g, 0.04 to 1.22 μg/g, and 2 to 25 ng/g, respectively. The predominant Fusarium species of the scabby wheat examined were F. sporotrichioides, F. avenaceum, F. poae, and F. crookwellense. Fifteen of 48 F. poae isolates and all four F. crookwellense isolates were screened for the production of seven derivatives of trichothecenes and zearalenone respectively, on rice culture. One isolate of F. poae produced diacetoxyscirpenol alone (4.3 μg/g); seven produced nivalenol (1.3 to 23.8 μg/g), 4-acetylnivalenol (0.1 to 4.6 μg/g), and diacetoxyscirpenol (0.9 to 99.5 μg/g); and five produced nivalenol alone (0.4 to 3.5 μg/g). The remaining two isolates produced no trichothecenes. Zearalenone production was not found in any isolate of F. poae tested. All isolates of F. crookwellense produced nivalenol (0.9 to 22.5 μg/g), 4-acetylnivalenol (0.5 to 25.0 μg/g), and zearalenone (1.4 to 162.5 μg/g). From these results, it is apparent that deoxynivalenol and zearalenone, and occasionally nivalenol, occur naturally throughout Hokkaido, and it is suggested that nivalenol-producing F. poae and F. crookwellense strains are responsible for the natural contamination with nivalenol found in the northernmost area of Japan. Furthermore, it was found for the first time that several isolates of F. poae distributed in Hokkaido possessed the ability to produce both type A and type B trichothecenes.     

cis-Elements and Protein Factors Related to Regulation of Transcription of Arylsulfatase Gene during Sea Urchin Development

Eight restriction fragments (I–VIII) were prepared to cover a whole span of the enhancer region in the upstream of the Ars gene of the sea urchin, Hemicentrotus pulcherrimus , and their abilities to influence on the Ars gene expression were estimated by CAT assay. Only three fragments (III, IV and V) encompassing a 0.6 kb region between −2.8 kb and −2.2 kb stimulated CAT expression. By mobility shift assays, it was found that the Ars enhancer region is composed of multiple cis -acting elements that interact with nuclear proteins in a sequence-specific manner. Among them, two sequences, a G-string and a GATCTCCCC, were determined by DNA footprinting as sites of protein-DNA interaction. The DNA-binding factor prevalence changed ontogenically in three different patterns. Possible activation of DNA-binding proteins through their modification is discussed.     

One-dimensional food chain equations and their generalization

Two equations describing one-dimensional food chains are known to possess soliton solutions. It is demonstrated that both equations are embraced within another equation, which arises in the theory of chains of enzymic reactions. We find an elliptic function solution to this equation. We obtain a one-soliton solution from it and re-derive the elliptic function solutions of the two ecological equations.     

Effect of the sugar moiety on complex formation of cycloamyloses with p-nitrophenyl glycosides

Circular diochroism (CD) spectra of four p-nitrophenyl glycosides and their cycloamylose complexes were investigated at various concentrations of cycloamylose and at temperatures ranging from 20 to 60°C. The CD spectra of p-nitrophenyl glycosides changed remarkably in the presence of cycloamyloses, and significant differences in spectral shape and intensity were observed between the cyclohexaamylose complex and the cycloheptaamylose complex. The difference CD spectra between the free guest and its complex indicates that the nitrophenyl group is included in the cycloamylose cavity but its disposition is different between the complexes with cyclohexaamylose and cycloheptaamylose. Values of enthalpy and entropy of the cyclohexaamylose complex are considerably larger than those of the corresponding cycloheptaamylose complex, although the free energy differs only slightly. It is suggested that the nitrophenyl group is more loosely bound to the cycloheptaamylose cavity than to the cyclohexaamylose cavity, and has much more flexibility in its disposition.