Immunochemical characterisation and epitope mapping of a novel fimbrial protein (Pg-II fimbria) of Porphyromonas gingivalis

Abstract Mouse monoclonal antibody (mAb) Pgf-II specific for a 72-kDa major cell-surface protein (72K-CSP) derived from Porphyromonas gingivalis OMZ 409 was prepared. Immunoblotting analysis revealed that mAb Pgf-II reacted with 72K-CSP but not with 41-kDa fimbrial subunit protein (41K-fimbrilin) derived from P. gingivalis 381. Electron microscopic observation revealed that P. gingivalis OMZ 409 possessed peritrichous, thin fimbriae on their surface. Immunogold electron microscopy also demonstrated that mAb Pgf-II bound to the 72K-CSP examined with the gold particles arranged along the fibril array originating from the cell surface of the bacteria. These findings suggested that P. gingivalis 72K-CSP was identifiable as another fimbriae (termed Pg-II fimbriae) different from the fimbriae (termed Pg-I fimbriae) composed of a 41K-fimbrilin. Using multipin peptide synthesis technology, 102 sequential overlapping peptides covering the entire 514 amino-acid stretch of Pg-II fimbriae were synthesised. Seven immunodominant regions within Pg-II fimbrial protein molecule, which definitely reacted with the serum of patients with periodontal diseases, were detected.     

Clonal propagation of Stevia rebaudiana Bertoni by stem-tip culture

Clonal propagation of Stevia rebaudiana has been established by culturing stem-tips with a few leaf primordia on an agar medium supplemented with a high concentration (10 mg/l) of kinetin. Anatomical examination has suggested that these multiple shoots originate from a number of adventitious buds formed on the margin of the leaf. Innumerable shoots can be obtained by repeating the cycle of multiple-shoot formation from a single stem-tip of Stevia. These shoots produce roots when transferred to a medium containing NAA (0.1 mg/l) without kinetin. The regenerated plantlets can be transplanted to soil.     

Absorption of Copper by Lemna as Influenced by Some Factors Which Nullify the Copper Effect on Flowering and Growth

The effect of copper on flowering and growth of Lemna paucicostata6746 and Lemna gibba G3 in a copper-containing medium is nullifiedby the addition of EDTA, ammonium ions or salicylic acid tothe medium or a decrease in its nitrate concentration. Thesefactors were examined for their effects on the absorption ofcopper by the plants. The addition of EDTA to the medium completelyinhibited the absorption of copper in both species, thus eliminatingthe copper effect. Ammonium ions also inhibited copper absorption,their effectiveness rising with their concentration. Loweringthe nitrate concentration in the medium nullified the coppereffect on flowering in L. paucicostata 6746, and the additionof salicylic acid to the medium also nullified the copper effectin L. gibba G3, both without affecting the absorption of copper. (Received June 7, 1982; Accepted August 27, 1982)     

Population dynamics of the swallowtail butterfly,Papilio xuthus L., in a deforested area

The long-term population dynamics of the swallowtail butterfly, Papilio xuthus, were studied by means of life table analysis in a deforested area, where the host tree, Zanthoxylum ailanthoides, was growing. The number of eggs laid on host trees in the deforested area decreases as secondary succession progresses. When the trees were classified into three groups, i. e. tall, medium and short, according to their height relative to the surrounding vegetation, less eggs were laid on tall than on short host trees. Life tables for the natural populations in each generation were developed on the basis of mean values for 6 years. Eggs and larvae in early stages were attacked chiefly by small- and middle-sized predators, such as ants, spiders, bugs and orthopterids, while later stage larvae were attacked chiefly by birds or Polistes wasps. An inchneumonid wasp was the most important mortality factor in the pupal stage. Key-factor analysis was tested on the life table data of each immature stage. The late larval stage was a key stage, and most mortality factors were considered to operate independently. The test for density dependency did not show any tendencies. The analysis of variance in two-way classifications was carried out for the difference of survival rate among the tree groups. It was suggested that tall host trees were unsuitable for P. xuthus. The construction of life tables for artificially inoculated populations on large host trees showed that high physiological death rate were characteristic of such trees in comparison with natural populations on smaller trees. The characteristics of the population dynamics of P. xuthus in the deforested area are compared among some kinds of the habitat, and the interrelationship between P. xuthus and its host plant are discussed.     

Hydrogen exchange kinetics of nucleic acids: Double and triple helices with Hoogsteen-type basepairs

The kinetics of hydrogen-tritium exchange reaction have been followed by a Sephadex technique of a double-helical poly(ribo-2-methylthio-adenylic acid)·poly(ribouridylic acid) complex with the Hoogsteen-type basepair. Only one hydrogen in every 2-methylthio-adenine·uracil basepair has been found to exchange at a measurably slow rate, 0.023 s^?1 (at 0°C), which is, however, much greater than that for a double-helix with the Watson-Crick type A·U pair. The kinetics of hydrogen-tritium exchange were also examined by triple-helical poly(rU)·poly(rA)·poly(rU) which involves both the Watson-Crick and Hoogsteen basepairings. Here, three hydrogens in every U·A·U base triplet have been found to exchange at a relatively slow rate, 0.0116 s^?1 (at 0°C). The kinetics of hydrogen-deuterium exchange reactions of these polynucleotide helices have also been followed by a stopped-flow ultraviolet absorption spectrophotometry at various temperatures. On the basis of these experimental results, the mechanism of the hydrogen exchange reactions in these helical polynucleotides was discussed. In the triple helix, the rate-determining process of the slow exchange of the three (one uracil-imide and two adenine-amino) hydrogens is considered to be the opening of the Watson-Crick part of the U·A·U triplet. This opening is considered to take place only after the opening of the Hoogsteen part of the triplet.     

Translocation of colicin E1 through cytoplasmic membrane of Escherichia coli

The product of the malE—lacZ gene fusion was reported to compete with some proteins including outer membrane lipoprotein in the protein translocation across the Echerichia coli membrane. The fusion product also inhibited colicin E1 export. Furthermore, globomycin, which accumulated prolipoprotein in the membrane, inhibited the translocation of colicin E1 in the wild-type cells, but not in lipoprotein-negative mutant cells. Since colicin E1 contains the internal signal-like sequence [Proc. Natl. Acad. Sci. USA (1982) 79, 2827–2831], these results suggest that colicin E1 is exported by the aid of this sequence at a common site for maltose-binding protein and lipoprotein translocation.     

Optical measurements of intracellular oxygen concentration of rat heart in vitro

The optical characteristics of hemoglobin-free perfused rat heart have been examined in detail. Ethyl hydrogen peroxide is found to convert myoglobin into “ferryl compound” in the perfused heart, as is also seen in vitro. After pretreatment with ethyl hydrogen peroxide, a typical mitochondrial absorption spectrum, similar to that of isolated rat heart mitochondria, is obtained in perfused heart. The overall absorption spectrum of the heart obtained by the aerobic to anaerobic transition is a superposition of the mitochondrial spectrum on that of myoglobin. By comparing these spectra, it is found that measurement of cytochrome a + a₃ at 605–620 nm is possible in spite of the absorbance change due to the oxygenation-deoxygenation of myoglobin, whereas the wavelength pairs for cytochrome c at 550-540 nm, cytochrome b at 562–575 nm and cytochrome a + a₃ at 445–450 nm can not be used in the heart because of interference from the absorption change of myoglobin. The partial pressure of O₂ (P₅₀) which is required for half maximal deoxygenation (or oxygenation) of myoglobin in perfused heart is found to be 2.4 mm Hg at room temperature and the Hill constant, n, is 1.1; these values are similar to those of myoglobin purified from rat heart. The steady-state O₂ titration has been performed by using absorbancy changes of myoglobin and cytochrome a + a₃ as intracellular O₂ indicators. In the perfused heart, the percentage change of oxygenation-deoxygenation of myoglobin parallels the oxidation-reduction of cytochrome a + a₃, while the mixture of purified myoglobin and isolated mitochondria shows a deviation, reflecting the difference of O₂ affinities between myoglobin and cytochrome a + a₃. The results indicate that there may be an O₂ gradient between cytosolic and mitochondrial compartments in the hemoglobin-free perfused heart. The absorption changes of myoglobin and of cytochrome a + a₃ can be measured in a single contraction-relaxation cycle. A triple beam method was introduced to eliminate the effect of light scattering changes in these measurements. The results demonstrated that myoglobin is more oxygenated during the systolic and diastolic periods and deoxygenated in the resting period, whereas cytochrome a + a₃ is more reduced in systole and diastole and oxidized in the resting state. Changing the perfusion conditions greatly alters the time course of the events which occur during the contraction-relaxation cycle of the perfused heart.