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1.
Summary A hybrid plasmid, pOXN29 (10.4 Mdal), coding the xylanase (xynA) and -xylosidase (xynB) genes of Bacillus pumilus IPO was constructed by the ligation of pBR322 and a 7.7 Mdal PstI-fragment of chromosomal DNA as reported in our previous paper (Panbangred et al. 1983). A deletion plasmid of pOXN29, pOXN293 (9.2 Mdal), which contains xynA and xynB, was ligated with pUB110 at an EcoRI site, and used to transform B. subtilis MI111. Two selected clones of B. subtilis as xylanase hyper-producers contained plasmids pOXW11 (4.2 Mdal) and pOXW12 (4.0 Mdal), both consisting of only pUB110, xynA, and its flanking regions, as the result of spontaneous deletion. These B. subtilis clones produced 2.7–3.0 times as much xylanase as B. pumilus. Escherichia coli and B. subtilis clones harbouring the hybrid plasmids synthesized xylanase and -xylosidase constitutively, whereas both enzymes were induced by xylose in B. pumilus.Xylanase synthesized by B. subtilis harbouring pOXW11 or pOXW12 was excreted into the medium like that of B. pumilus IPO, but xylanase synthesized in E. coli harbouring pOXN29, 293 or pOXW1 coding xynA was intracellular. In a previous investigation (Panbangred et al. 1983), xylanase was found to be located in the cytoplasm, not the periplasm nor the membrane fraction in E. coli cells harbouring pOXN29 derivatives. In spite of the abnormal location of xylanase synthesized in E. coli, the signal peptide was processed in the same way as in B. pumilus, with the same molecular weight and the same amino terminal sequences of xylanase prepared from E. coli cells and B. pumilus culture fluid.  相似文献   
2.
Serum-free culture of rat keratinocytes   总被引:2,自引:0,他引:2  
Summary Procedures for the serum-free culture of rat keratinocytes have been established. Basal cells prepared from epidermis of newborn rat were stored in liquid nitrogen and used for primary culture. Among the available media, MCDB 153, developed originally for human keratinocyte (HK) culture, was the best for the development of serum-free formulation. To grow rat keratinocytes, bovine serum albumin was arbitrarily substituted for the macromolecule supplements needed for HK culture, i.e. fetal bovine serum protein or bovine pituitary extract. Qualitative and quantitative adjustment of supplements was thereafter made to support rapid cell growth. Satisfactory cell growth was achieved in the optimized medium of MCDB 153 supplemented with growth factors and amino acids: insulin (10 μg/ml), hydrocortisone (0.1 μg/ml), epidermal growth factor (25 ng/ml), calcium chloride (0.2 mM), histidine (0.23 mM), isoleucine (0.05 mM), tryptophane (0.015 mM), threonine (1.25 mM), tyrosine (0.031 mM), alanine (4.08 mM), and albumin (2 mg/ml). This optimized culture system was superior to the original HK culture condition for rapid growth of rat keratinocytes. Under our condition, cells grew as a monolayer, becoming confluent, but without stratification, and were passaged 2 to 3 times without any changes in morphology. The serum-free formulation allows us to control more accurately the concentrations of biomolecules in the medium including lipids and hormones, and therefore will be suitable for the study focusing on lipid metabolism or hormonal regulation of rat keratinocytes.  相似文献   
3.
4.
The structure for trichoclin, (E)-8-(3-methyl-4-hydroxy-2-butenyloxy)-psoralen, a new furocoumarin isolated from Trichocline incana, has been established. Phellopterin and isopimpinellin were also obtained. The new side chain of trichoclin was confirmed by synthesis.  相似文献   
5.
Summary The regulatory mechanism of ATP regeneration by the glycolytic pathway in Hansenula jadinii cells was investigated by analyzing the initial stage of CDP-choline fermentation. As a result, the on-off of ATP regeneration was found to be determined by the ATP concentration overcoming the inhibitory effect of phosphate buffer on hexokinase activity. The concentration of ATP at the initial stage of fermentation was greatly influenced by the kinds and amounts of glycogen in cells. Based on these results, the regulatory mechanism of ATP regeneration by the glycolytic pathway is discussed in detail.  相似文献   
6.
Oxidative Degradation of Squalene by Arthrobacter Species   总被引:5,自引:5,他引:0       下载免费PDF全文
An organism isolated from soil and identified as Arthrobacter sp. was studied for its squalene degradation. The degradation product from squalene, which accumulated in the culture broth, was isolated and identified as trans-geranylacetone by mass spectrometry, gas chromatography, infrared spectrometry, and nuclear magnetic resonance spectrometry. Addition of a high concentration of K2HPO4 to the culture medium resulted in accumulation of fairly large amounts of carboxylic acids in addition to geranylacetone. These carboxylic acids were identified as isovaleric, β,β′-dimethylacrylic, geranic, and (+)-(R)-citronellic acids. Among these acids, α,β-saturated carboxylic acids were found to be predominant in quantity.  相似文献   
7.
Human gammadelta T cells display unique repertoires of Ag specificities largely imposed by selective usages of distinct Vgamma and Vdelta genes. Among them, Vgamma2/Vdelta2(+) T cells predominate in the circulation of healthy adults and respond to various microbial small molecular mass nonpeptide Ags. The present results indicate that the primary Vgamma2/Vdelta2(+) T cells stimulated with the distinct groups of nonpeptide Ags, including monoethyl pyrophosphate, isobutyl amine, and aminobisphosphonate, invariably exhibit Jgamma1.2 in the Vgamma2(+) TCR-gamma chains. Gene transfer studies revealed that most of the randomly cloned Vgamma2/Jgamma1.2(+) TCR-gamma genes bearing diverse Vgamma/Jgamma junctional sequences could confer the responsiveness to all these nonpeptide Ags, while none of the Vgamma2/Jgamma1.1(+) or Vgamma2/Jgamma1.3(+) TCR-gamma genes could do so. Furthermore, mutation of the lysine residues encoded by the Jgamma1.2 gene, which are unique in human Jgamma1.2 and absent in other human or mouse Jgamma segments, completely abrogated the responsiveness to all the nonpeptide Ags without affecting the response to anti-CD3 mAb. These results strongly suggested that the positively charged lysine residues in the TCR-gamma chain CDR3 region encoded by the germline Jgamma1.2 gene play a key role in the recognition of diverse small molecular mass nonpeptide Ags.  相似文献   
8.
Since S-n-butyl S′-p-tert-butylbenzyl N-3-pyridyldithiocarbonimidate, a potent fungicide to powdery mildew, is known to inhibit ergosterol biosynthesis in Monilinia fructigena, the activities were assessed on 24 compounds having other substituents than the 3-pyridyl and on 24 compounds having a variety of different structures connecting the 3-pyridyl and the p-tert-butylphenyl group from that of the dithiocarbonimidate.

In the former group the 3-pyridyl group was essential for the activities and the substitution at the 2- or 6-position resulted, on available data, in inactive compounds. Several other β-N-heterocyclic analogs were also active. In the latter group, a number of modified compounds from the dithiocarbonimidate structure were shown to be active.  相似文献   
9.
Journal of Plant Research - Cysteine biosynthesis is directed by the successive commitments of serine acetyltransferase, and O-acetylserine (thiol) lyase (OASTL) compounds, which subsequently frame...  相似文献   
10.
Prostaglandins are biologically active substances used in a wide range of medical treatments. Prostaglandins have been supplied mainly by chemical synthesis; nevertheless, the high cost of prostaglandin production remains a factor. To lower the cost of prostaglandin production, we attempted to produce prostaglandins using a liverwort, Marchantia polymorpha L., which accumulates arachidonic acid, which is known as a substrate of prostaglandins. Here we report the first bioproduction of prostaglandins in plant species by introducing a cyclooxygenase gene from a red alga, Gracilaria vermiculophylla into the liverwort. The transgenic liverworts accumulated prostaglandin F, prostaglandin E2 and prostaglandin D2 which were not detected in the wild-type liverwort. Moreover, we succeeded in drastically increasing the bioproduction of prostaglandins using an in vitro reaction system with the extracts of transgenic liverworts.  相似文献   
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