Stability of plasmid PBR322 in Escherichia coli immobilized on cotton cloth during use as resident inoculum

Summary E. coli cells harbouring plasmid pBR322 which confers ampicillin resistance were immobilized on cotton cloth. The resulting film was used as an inoculum in daily repeated batch culture in ampicillin-free medium. During two months, the film was able to produce cultures which, at the late log phase, showed little sensitivity to 10 mg/ml ampicillin. Thus such a bacterial film can effectively be used as an inoculum for the production of recombinant DNA products by means of pBR322 or its derivatives in the absence of ampicillin.     

Biosynthesis of Purine Alkaloids in Camellia Plants

The metabolism of [8-¹⁴C]adenine and [8-¹⁴C]hypoxanthine infour species of Camellia plants was investigated in relationto the synthesis of purine alkaloids, caffeine and theobromine.Young leaves of C. sinensis had the ability to synthesize caffeine,but in C. irrawadiensis, these labelled precursors were incorporatedinto theobromine, not caffeine. No synthesis of purine alkaloidscould be detected in C. japonica and C. sasanqua leaves. Conventional"salvage" and degradation pathways of purines were present inall species of Camellia plants examined. ¹ Present address: Research Center, Mitsubishi Chemical IndustriesLtd., 1000 Kamisida-cho, Midori-ku, Yokohama, 227 Japan. (Received September 29, 1986; Accepted January 22, 1987)     

Solubilization and Characterization of the Nitrendipine Receptor in Rat Brain

The nitrendipine receptor associated with the voltage-dependent calcium channel in rat brain was solubilized by detergent extraction and sonication. The detergent solution used for extraction consisted of 10 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), 0.25% (wt/vol) polyoxyethylene 20 cetyl ether (Brij 58), and 0.025% (wt/vol) polyoxyethylene 17 cetyl stearyl ether (Lubrol WX) in the presence of 30% (wt/vol) glycerol as a stabilizer. The molecular weight of the receptor was estimated to be 1,800K by Sephacryl S-500 gel filtration and 800K by sucrose density gradient sedimentation. The equilibrium dissociation constant of [3H]nitrendipine to the solubilized receptors was 5.6 nM, which is approximately 10 times that of the membrane-bound receptor. The binding of nitrendipine to the receptor was inhibited noncompetitively by the structurally unrelated calcium channel inhibitors verapamil and prenylamine; their concentrations for 50% inhibition were both 1.0 X 10(-7) M, and they caused maximal inhibitions of 70 and 100%, respectively.     

Taxonomic study of the genusMyagropsis (Cystoseiraceae,Phaeophyta)

The generic names of the brown algaeMyagropsis, Cystoseira andCystophyllum have been used in Japan, Korea and China for a small group of seaweeds whose limits have not been clearly understood. Studies on the development of the eggs and subsequent germlings show that substantial differences occur betweenCystoseira andMyagropsis. Myagropsis Kützing is distinguished fromCystoseira C. Agardh by the following characteristics: (1) the tongue cell is undivided during development of the conceptacle; (2) paraphyses are projected from the conceptacle ostiole and become entangled; (3) during development, oospore germlings are mixed among paraphyses projecting from the ostiole; (4) oospores are large, with eight nuclei at maturity; (5) thirty-two primary rhizoids are produced on the germlings; and (6) the thallus is bilateral in organization. The shape and size of vesicles, their formation, and the presence of cryptostomata have been used as specific characters, but their use cannot be continued. It is concluded that the genusMyagropsis is monotypic, with a single species,M. myagroides (Turner) Fensholt. The status of this species is also discussed.     

Dynamics of appearance and disappearance of the ripple structure in multilamellar liposomes of dipalmitoylphosphatidylcholine

The physical properties of the pretransition (P beta'----L beta') of dipalmitoylphosphatidylcholine liposomes were investigated using freeze-fracture electron microscopy. The kinetics of pretransition examined in the previous paper using TEMPO spin probe (Tsuchida, K., et al. (1985) Biochim. Biophys. Acta 812, 249-254) was extensively studied by observing the ripple structures in the freeze-fractured surfaces at different time intervals. When the temperature is decreased from 38 degrees C to 30 degrees C, the ripple structure disappears in the following steps. The intervals between ripples begin to expand with the decrease of ripple density upon the temperature shift, and this process continues for several tens minutes. Then, each ripple disappears gradually and changes into a completely smooth surface at 3 h after the temperature shift. The comparison of relaxation times between the previous ESR measurement and the present experiment suggests that the fast relaxation observed in the previous study corresponds to the expansion of the intervals between ripples. On the other hand, the ripple structure of regular intervals appears rapidly in some places and then spreads over the whole area of fractured surface when the temperature is increased from 23 degrees C to 35 degrees C. The results obtained in this work and the previous ESR work strongly suggest that the formation and disappearance of ripple structure is closely related to the relaxation processes near the pretransition temperature.     

Postglacial environmental change of the Pacific Ocean off the coasts of central Japan

Downcore changes in microfossil assemblages and oxygen isotope ratios in three piston cores recovered from the Northwestern Pacific, off central Japan, show that the subtropical Kuroshio front was located to the south of C-4 core site (Lat. 33° N) during the last glacial. The front then advanced northward, passing over the C-4 site and the C-6 site (34.6° N) at about 13 ka and 10 ka, respectively, and reached the C-1 core site (36° N) at about 7 ka. After 5.5 ka it retreated to the area between the C-1 and C-6 sites. A brief but significant cold event, the readvance of the cold Oyashio Current, is recognized between 11 and 10 ka in the two northern cores, but the current did not reach the southern C-4 site. A contemporaneous cold event is known in the North Atlantic, and the cooling was probably a global phenomenon likely to be associated with lowering of sea level. Contamination of isotopically light water is apparent between 14 and 11 ka in the marked change in isotopic composition of benthic foraminifers. Oxygen isotope ratios of planktonic foraminifers show that prior to the advance of the Kuroshio front, the surface water at these core sites was isotopically lighter than the Kuroshio water at that time.     

Mutagenicity,sister chromatid exchange inducibility andin vitro cell transforming ability of particulates from Athens air

Airborne particulates were collected over a period of twelve months by the use of Hi-Vol samplers in the basin of Athens, Greece. N-Hexane extracts were tested in a battery ofin vitro tests for their ability to induce mutation in bacteria as well as mutation, sister chromatid exchange and morphological transformation in cultured mammalian cells. Positive results were found for mutagenicity withSalmonella strain TA98 in the Ames assay, for sister chromatid exchange induction in CHO cells and for transformation in BALB/c 3T3 cells in culture. They also showed weak non-doserelated induction of ouabain resistance in BALB/c 3T3 cells. The contribution of oxidizing and nitrating agents found in the Athens atmosphere, together with sunlight UV irradiation in the formation of direct acting mutagens and potential carcinogens from ambient polycyclic aromatic hydrocarbons, is suggested.Abbreviations FCS fetal calf serum - FPG fluorescent-plus-Giemsa technique - oua^R ouabain resistant - PAH polycyclic aromatic hydrocarbon - SCE sister chromatid exchange - TSP total suspended particulate     

Involvement in DNA repair of the ruvA gene of Escherichia coli

Summary The ruv operon of Escherichia coli consists of two genes, orfl1 and ruv, which encode 22 and 37 kilodalton proteins, respectively, and are regulated by the SOS system. Although the distal gene, ruv, is known to be involved in DNA repair, the function of orf1 has not been studied. To examine whether orf1 is also involved in DNA repair, we constructed a strain with a deletion of the entire ruv operon. The strain was sensitive to UV even after introduction of low copy number plasmids carrying either orf1 or ruv, but UV resistance was restored by introduction of a plasmid carrying both orfl and ruv. These results suggest that orf1 as well as ruv is involved in DNA repair. Therefore, orf1 and ruv should be renamed ruvA and ruvB, respectively.     

High levels of manganese-containing superoxide dismutase and thermally induced DNA disruption in a dnaK7(Ts) mutant of Escherichia coli K12

Summary IndnaK7(Ts) mutant cells, scission of DNA strands occurred after temperature shift up. When cells at 30°C were labeled with [³H]-thymidine and then shifted to 46° or 49°C for 20 min, the profiles of sedimentation of thier cellular DNA in an alkaline sucrose gradient revealed a decrease in the size of DNA to a quarter of that at 30°C in the mutant, but not in wild-type cells. The level of manganese-containing superoxide dismutase (MnSOD) in the mutant was about twice that in wild-type cells, even at the permissive temperature, implying increased production of superoxide radical anion, which may cleave DNA strands directly or indirectly in the mutant. Moderate increase in the MnSOD level on temperature shift up was observed in both strains. These results indicated that some components of the DnaK protein participate in protection of cellular membrane functions from thermal damage resulting from elevated production of the superoxide anion radical.     

Electron Transfer between Plastocyanin and P700 in Various Types of Photosystem I Complex

Three types of PS I Chl-protein complex, PS I 180, PS I 65,and PS I 30, have been prepared and the kinetic properties ofthe transfer of electrons from plastocyanin to P700 in the PSI complexes with different sized antennae were examined. ThePS I 180 complex, which consists of 180 Chi per P700, showedthe almost same rate constant and effects of cations for thetransfer of electrons from plastocyanin to P700 as those obtainedwith PS I-enriched membrane fragments. The rate constant increasedwith the addition of low concentrations of monovalent and divalentcations, but decreased with high concentrations of cations.However, the rate was severely reduced in the case of the PSI 65 and PS I 30 complexes, and quite different effects of cationswere observed. Given the presence of additional 25- to 28-kDapolypeptides in the PS I 180 complex as compared to the PS I65 and PS I 30 complexes, we discuss a possible function forthese polypeptides in the regulation of the reaction betweenplastocyanin and P700. ¹This work was supported in part by a Grant-in-Aid for ScientificResearch from the Ministry of Education, Science and Cultureof Japan. (Received May 27, 1988; Accepted November 7, 1988)