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1.
Hisashi Kato-Noguchi Madoka Yamamoto Kazuya Tamura Toshiaki Teruya Kiyotake Suenaga Yoshiharu Fujii 《Plant Growth Regulation》2010,60(2):127-131
Aqueous methanol extracts of rattail fescue (Vulpia myuros) inhibited the growth of roots and shoots of cress (Lepidium sativum), lettuce (Lactuca sativa), alfalfa (Medicago sativa), timothy (Phleum pratense), Digitaria sanguinalis and Lolium multiflorum. Increasing the extract concentration increased the inhibition, suggesting that rattail fescue may have growth inhibitory
substances and possess allelopathic potential. The aqueous methanol extract of rattail fescue was purified and two main inhibitory
substances were isolated and identified by spectral data as (−)-3-hydroxy-β-ionone and (+)-3-oxo-α-ionol. Both substances
inhibited root and shoot growth of cress at concentrations greater than 0.3 μM. The concentrations required for 50% growth
inhibition on root and shoot growth of cress, lettuce, alfalfa, timothy, D. sanguinalis and L. multiflorum were 2.7–19.7 μM for (−)-3-hydroxy-β-ionone, and 2.1–34.5 μM for (+)-3-oxo-α-ionol. The concentration of (−)-3-hydroxy-β-ionone
and (+)-3-oxo-α-ionol, respectively, in rattail fescue was 7.8 and 3.7 μg g−1 fresh weight. Considering the endogenous level and the inhibitory activity, (−)-3-hydroxy-β-ionone and (+)-3-oxo-α-ionol
may work as allelopathic substances in rattail fescue through the growth inhibition of neighboring plant species. 相似文献
2.
Yogan Khatri Marco Girhard Anna Romankiewicz Michael Ringle Frank Hannemann Vlada B. Urlacher Michael C. Hutter Rita Bernhardt 《Applied microbiology and biotechnology》2010,88(2):485-495
Sesquiterpenes are particularly interesting as flavorings and fragrances or as pharmaceuticals. Regio- or stereoselective
functionalizations of terpenes are one of the main goals of synthetic organic chemistry, which are possible through radical
reactions but are not selective enough to introduce the desired chiral alcohol function into those compounds. Cytochrome P450
monooxygenases are versatile biocatalysts and are capable of performing selective oxidations of organic molecules. We were
able to demonstrate that CYP109D1 from Sorangium cellulosum So ce56 functions as a biocatalyst for the highly regioselective hydroxylation of norisoprenoids, α- and β-ionone, which
are important aroma compounds of floral scents. The substrates α- and β-ionone were regioselectively hydroxylated to 3-hydroxy-α-ionone
and 4-hydroxy-β-ionone, respectively, which was confirmed by 1H NMR and 13C NMR. The results of docking α- and β-ionone into a homology model of CYP109D1 gave a rational explanation for the regio-selectivity
of the hydroxylation. Kinetic studies revealed that α- and β-ionone can be hydroxylated with nearly identical V
max and K
m values. This is the first comprehensive investigation of the regioselective hydroxylation of norisoprenoids by CYP109D1. 相似文献
3.
Wild-type cytochrome P450 monooxygenase from Bacillus megaterium (P450 BM-3) has a low hydroxylation activity for β-ionone (<1 min−1). Substitution of phenylalanine by valine at position 87 led to a more than 100-fold increase in β-ionone hydroxylation activity
(115 min−1). Enzyme activity could be further increased by both site-directed and random mutagenesis. The mutant R47L Y51F F87V, designed
by site-directed mutagenesis, and the mutant A74E F87V P386S, obtained after two rounds of error-prone polymerase chain reaction,
exhibited an increase in activity of up to 300-fold compared to the wild-type enzyme. The triple mutant R47 LY51F F87V exhibited
moderate enantioselectivity, forming (R)-4-hydroxy-β-ionone with an optical purity of 39%. All mutants regioselectively converted β-ionone into 4-hydroxy-β-ionone.
The regioselectivity is determined amongst others by the absolute configuration of the substrate. 相似文献
4.
Hiroshi Sakamaki Ken-ichi Itoh Wen Chai Yumiko Hayashida Susumu Kitanaka C. Akira Horiuchi 《Journal of Molecular Catalysis .B, Enzymatic》2004,27(4-6):177-181
Suspension cultures of Caragana chamlagu (Leguminosae) convert (±)-α-ionone (1) into (±)-3-oxo-α-ionone (3) as the major product and β-ionone (2) into 5,6-epoxy-β-ionone (6) as the sole product. It is interesting to note that the cultured cells of C. chamlagu convert regioselectively the cycloolefinic part of 1 into the corresponding unsaturated carbonyl compound, allylic alcohol and epoxide as the oxidation products, whereas the suspension cultures of Nicotiana tabacum (Solanaceae) convert the unsaturated carbonyl of 1 into the corresponding saturated ketones and alcohols as reduction products. 相似文献
5.
Cavalieri D Casalone E Bendoni B Fia G Polsinelli M Barberio C 《Molecular & general genetics : MGG》1999,261(1):152-160
Seven spontaneous Saccharomyces cerevisiae mutants that express dominant resistance to 5,5,5-trifluoro-DL-leucine have been characterised at the molecular level. The
gene responsible for the resistance was cloned from one of the mutants (FSC2.4). Determination of its nucleotide sequence
showed that it was an allele of LEU4 (LEU4-1), the gene that encodes α-isopropyl malate synthase I (α-IPM synthase I), and that the mutation involved a codon deletion
localised close to the 3′ end of the LEU4 ORF. Six different point mutations – four transitions and two transversions – were found in the remaining mutants. α-IPM
synthase activity was found to be insensitive to feedback inhibition by leucine in five of the strains. In the other two the
enzyme was resistant to Zn2+-mediated inactivation by Coenzyme A, a previously postulated control mechanism in energy metabolism; as far as we know, this
represents the first direct in vivo evidence for this mechanism. The seven mutations define a region, the R-region, involved
in both leucine feedback inhibition and in Zn2+-mediated inactivation by CoA. Deletion experiments involving the R-region showed that it is also necessary for enzyme activity.
Received: 30 September 1998 / Accepted: 20 October 1998 相似文献
6.
Bleif S Hannemann F Zapp J Hartmann D Jauch J Bernhardt R 《Applied microbiology and biotechnology》2012,93(3):1135-1146
The use of cytochromes P450 for the regio- and stereoselective hydroxylation of non-activated carbon atoms in biotechnological
applications reflects an efficient and cost-effective alternative in comparison to classical organic chemistry. The prokaryotic
cytochrome P450 CYP106A2 from Bacillus megaterium ATCC 13368 hydroxylates a variety of 3-oxo-Δ4 steroids and recently it was identified to carry out a one-step regioselective allylic hydroxylation of the diterpene abietic
acid. The anti-inflammatory pentacyclic triterpene 11-Keto-β-boswellic acid (KBA) was found to be a further substrate of CYP106A2,
being the first report of a pentacyclic triterpene conversion by a prokaryotic P450. The reaction products were analyzed by
HPLC and the corresponding kinetic parameters were investigated. Structure determination of the main product by NMR revealed
a 15α-hydroxylation of this substrate. In order to overcome the inability of a recombinant P450 whole-cell system in E. coli for the uptake of acids with terpene structure, we developed for the first time an expression system for cytochromes P450
in B. megaterium (strains MS941 and ATCC 13368). Interestingly, CYP106A2 was only successfully expressed in the plasmid-less B. megaterium strain MS941 but not in ATCC13368. This recombinant system, with the co-expressed heterologous redox chain of the P450, bovine
adrenodoxin reductase (AdR), and bovine adrenodoxin (Adx), was applied for the whole-cell conversion of KBA. The formation
of 15α-hydroxy-KBA was increased 15-fold in comparison with the naturally CYP106A2-expressing B. megaterium strain ATCC 13368. 相似文献
7.
The cloning of α-amylase gene ofS. occidentalis and the construction of starch digestible strain of yeast,S. cerevisiae AS. 2. 1364 with ethanol-tolerance and without auxotrophic markers used in fermentation industry were studied. The yeast/E.coli shuttle plasmid YCEp1 partial library ofS. occidentalis DNA was constructed and α-amylase gene was screened in S.cerevisiae by amylolytic activity. Several transformants with amylolysis were obtained and one of the fusion plasmids had an about 5.0
kb inserted DNA fragment, containing the upstream and downstream sequences of α-amylase gene fromS. occidentalis. It was further confirmed by PCR and sequence determination that this 5.0 kb DNA fragment contains the whole coding sequence
of α-amylase. The amylolytic test showed that when this transformant was incubated on plate of YPDS medium containing 1 %
glum and 1 % starch at 30°C for 48 h starch degradation zones could be visualized by staining with iodine vapour. α-amylase
activity of the culture filtratate is 740–780 mU/mL and PAGE shows that the yeast harboring fusion plasmids efficiently secreted
α-amylase into the medium, and the amount of the recombinant α-amylase is more than 12% of the total proteins in the culture
filtrate. These results showed that α-amylase gene can be highly expressed and efficiently secreted inS. cerevisiae AS. 2.1364, and the promotor and the terminator of α-amylase gene fromS. occidentalis work well inS. cercvisiac AS. 2.1364. 相似文献
8.
Bentel JM Pickering MA Pollard M Clements JA Tilley WD 《In vitro cellular & developmental biology. Animal》1999,35(10):655-662
Summary Prostate tumors were induced in Lobund-Wistar rats by treatment with N-methyl-N-nitrosourea (MNU) and testosterone propionate (TP). Androgen receptor (AR) expression was confirmed in 16 (100%) of the primary
prostate cancers, with strong uniform staining in well-differentiated tumors and more variable AR immunoreactivity in poorly
differentiated tumors. Epithelial cell lines were established from nine of the tumors. At early passages, four of the tumor
cell lines tested were strongly immunoreactive for AR; however, only two of the cell lines, E2(A) and F2, have remained AR-positive.
These cell lines specifically bind 3H-DHT at 40 and 19 fmol/mg protein, respectively, and express a 110 kDa AR immunoreactive protein. Proliferation in in vitro
culture of both E2(A) and F2 cells was increased in the presence of 5α-dihydrotestosterone (DHT). The antiandrogen, hydroxyflutamide was able to prevent the DHT-induced growth of E2(A) but not
F2 cells. Furthermore, hydroxyflutamide alone increased proliferation of F2 cells, suggesting that the androgen signalling
pathway in this cell line may be abnormal. Tumorigenicity of the AR-expressing and nonexpressing cell lines was confirmed
by xenograft formation following subcutaneous inoculation into intact male nude mice. In summary, carcinogen-induced prostate
tumors of Lobund-Wistar rats express AR and two of nine cell lines derived from the tumors express AR. Further evaluation
of AR structure in primary prostate tumors forming spontaneously or following MNU and TP induction will determine whether,
as in human prostate cancers, disease progression in Lobund-Wistar rats is associated with mutations in the AR gene. 相似文献
9.
Hiroyuki Uehara Bok Du Choi Enoch Y. Park Mitsuyasu Okabe 《Biotechnology and Bioprocess Engineering》2000,5(1):7-12
The expression of the mouse α-amylase gene in the methylotrophic yeast,P. pastoris was investigated. The mouse α-amylase gene was inserted into the multi-cloning site of a Pichia expression vector, pPIC9,
yielding a new expression vector pME624. The plasmid pME624 was digested withSalI orBglII, and was introduced intoP. pastoris strain GS115 by the PEG1000 method. Fifty-three transformants were obtained by the transplacement of pME624 digested withSalI orBglII into theHIS
4 locus (38 of Mut+ clone) or into theAOX1 locus (45 of Muts clone). Southern blot was carried out in 11 transformants, which showed that the mouse α-amylase gene was integrated into
thePichia chromosome. When the second screening was performed in shaker culture, transformant G2 showed the highest α-amylase activity,
290 units/ml after 3-day culture, among 53 transformants. When this expression level of the mouse α-amylase gene is compared
with that in recombinantSaccharomyces cerevisiae harboring a plasmid encoding the same mouse α-amylase gene, the specific enzyme activity is eight fold higher than that of
the recombinantS. cerevisiae. 相似文献
10.
Wei-Wei Zhang Xiao-Juan Duan Hai-Lan Huang Yi Zhang Bin-Gui Wang 《Journal of applied phycology》2007,19(2):97-108
The extracts obtained from 28 species of marine algae were evaluated for their antioxidant activity (AA) versus the positive
controls butylated hydroxytoluene (BHT), gallic acid (GA), and ascorbic acid (AscA). Most of the tested samples displayed
antioxidant activity to various degrees. Among them, the extract of Symphyocladia latiuscula exhibited the strongest AA, which was comparable to BHT, GA, and AscA in radical scavenging activity, as shown in the DPPH
(α,α-diphenyl-β-picrylhydrazyl) assay, and higher than those of the positive controls in β-carotene-linoleate assay system.
In addition, the ethyl acetate-soluble fraction isolated from the crude extract of S. latiuscula exhibited the highest antioxidant activity in both assay systems. This fraction was further fractionated into seven subfractions
(F1-F7) by vacuum liquid chromatography (VLC). F1 and F4 were found to be the most effective subfractions in scavenging DPPH
radical assay and in the β-carotene-linoleate assay, respectively. The total phenolic content (TPC) and reducing power (RP)
for all of the extracts, fractions, and subfractions (F1–F7) were also determined. The TPC of the 28 extracts ranged from
0.10 to 8.00 gallic acid equivalents (mg/g seaweed dry weight) while the RP ranged from 0.07 to 11.60 ascorbic acid equivalents
(mg·g−1 seaweed dry weight). Highly positive relationships between AA and TPC as well as between AA and RP were found for the extracts
and fractions, while for the subfractions F1–F7 only weak or no such relations were found. The results obtained from this
study indicate that further analysis is needed of those marine algal species that contain the most antioxidant activity in
order to identify the active principles. 相似文献
11.
Murai T Ueda M Shibasaki Y Kamasawa N Osumi M Imanaka T Tanaka A 《Applied microbiology and biotechnology》1999,51(1):65-70
The construction of a whole-cell biocatalyst with its sequential reaction has been performed by the genetic immobilization
of two amylolytic enzymes on the yeast cell surface. A recombinant strain of Saccharomyces cerevisiae that displays glucoamylase and α-amylase on its cell surface was constructed and its starch-utilizing ability was evaluated.
The gene encoding Rhizopus oryzae glucoamylase, with its own secretion signal peptide, and a truncated fragment of the α-amylase gene from Bacillus stearothermophilus with the prepro secretion signal sequence of the yeast α factor, respectively, were fused with the gene encoding the C-terminal
half of the yeast α-agglutinin. The constructed fusion genes were introduced into the different loci of chromosomes of S. cerevisiae and expressed under the control of the glyceraldehyde-3-phosphate dehydrogenase promoter. The glucoamylase and α-amylase
activities were not detected in the culture medium, but in the cell pellet fraction. The transformant strain co-displaying
glucoamylase and α-amylase could grow faster on starch as the sole carbon source than the transformant strain displaying only
glucoamylase.
Received: 16 June 1998 / Received last revision: 21 August 1998 / Accepted: 3 September 1998 相似文献
12.
Summary Replacement of the regulatory sequence of theBacillus amyloliquefaciens α-amylase gene (AMY1) by the yeast alcohol dehydrogenase gene promoter (ADC1
p) resulted in increased levels of extracellular α-amylase production inSaccharomyces cerevisiae. Negative regulation of glucoamylase synthesis by theSTA10-encoded repressor was alleviated by replacing the nativeSTA2 gene promoter fromS. cerevisiae var.diastaticus withADC1
p. Enhanced degradation of starch was achieved when the modified versions of theAMY1 andSTA2 genes were introduced jointly intoS. cerevisiae. 相似文献
13.
A. V. Perepelov Bin Liu S. N. Senchenkova A. S. Shashkov S. D. Shevelev Lu Feng Lei Wang Y. A. Knirel 《Biochemistry. Biokhimii?a》2010,75(1):19-24
On mild acid degradation of the lipopolysaccharide of Escherichia coli O108, the O-polysaccharide was isolated and studied by sugar analysis and one- and two-dimensional 1H- and 13C-NMR spectroscopy. The polysaccharide was found to contain an unusual higher sugar, 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-d-galacto-non-2-ulosonic acid (di-N-acetyl-8-epilegionaminic acid, 8eLeg5Ac7Ac). The following structure of the tetrasaccharide repeating unit of the polysac-charide
was established: →4)-α-8eLegp5Ac7Ac-(2→6)-α-D-Galp-(1→3)-α-L-FucpNAc-(1→3)-α-D-GlcpNAc-(1→. Functions of the E. coli O108 antigen biosynthetic genes, including seven putative genes for synthesis of 8eLeg5Ac7Ac, were assigned by sequencing
the O-antigen gene cluster along with comparison with gene databases and known biosynthetic pathways for related nonulosonic
acids. 相似文献
14.
15.
Yoshinari Yamazaki Yumiko Hayashi Masatoshi Arita Tadaharu Hieda Yoichi Mikami 《Applied microbiology》1988,54(10):2354-2360
α-Ionone, α-methylionone, and α-isomethylionone were converted by Aspergillus niger JTS 191. The individual bioconversion products from α-ionone were isolated and identified by spectrometry and organic synthesis. The major products were cis-3-hydroxy-α-ionone, trans-3-hydroxy-α-ionone, and 3-oxo-α-ionone. 2,3-Dehydro-α-ionone, 3,4-dehydro-β-ionone, and 1-(6,6-dimethyl-2-methylene-3-cyclohexenyl)-buten-3-one were also identified. Analogous bioconversion products from α-methylionone and α-isomethylionone were also identified. From results of gas-liquid chromatographic analysis during the fermentation, we propose a metabolic pathway for α-ionones and elucidation of stereochemical features of the bioconversion. 相似文献
16.
Bacillus pumilis F3-4 utilized feather as a sole source of carbon, nitrogen and sulfur. Supplementation of the feather medium with glucose
or MgSO4 · 7H2O increased keratinolytic protease production (14.6–16.7 U/mg). The synthesis of keratinolytic protease was repressed by an
exogenous nitrogen source. Keratinolytic protease was produced in the absence of feather (9.4 U/mg). Feather degradation resulted
in sulfhydryl group formation (0.8–2.6 μM). B. pumilis F3-4 effectively degraded chicken feather (75%), duck feather (81%) and feather meal (97%), whereas human nails, human hair
and sheep wool under went less degradation (9–15%).
An erratum to this article can be found at 相似文献
17.
Cira LA González GA Torres JC Pelayo C Gutiérrez M Ramírez J 《Antonie van Leeuwenhoek》2008,93(3):259-266
This paper describes the effect of the heterologous expression of tomatinase from Fusarium oxysporum f. sp lycopersici in Saccharomyces cerevisiae. The gene FoTom1 under the control of the S. cerevisiae phosphoglycerate kinase (PGK1) promoter was cloned into pYES2. S. cerevisiae strain Y45 was transformed with this vector and URA3 transformant strains were selected for resistance to α-tomatine. Two
transformants were randomly selected for further study (designated Y45-1 and Y45-2). Control strain Y45 was inhibited at 50 μM
α-tomatine, in contrast, transformants Y45-1 and Y45-2 did not show inhibition at 200 μM. Tomatinase activity was detected
by HPLC monitoring tomatine disappearance and tomatidine appearance in the supernatants of culture medium. Maximum tomatinase
activity was observed in the transformants after 6 h, remaining constant during the following 24 h. No tomatinase activity
was detected in the parental strain. Moreover, the transformants were able to grow and produce ethanol in a mix of Agave
tequilana Weber var. azul and Agave salmiana must, contrary to the Y45 strain which was unable to grow and ferment under these conditions. 相似文献
18.
The micronuclear genes encoding α-telomere-binding protein (αTP) in Oxytricha trifallax and Stylonychia mytilus contain multiple internal eliminated segments, or IESs, that divide the gene into multiple parts called macronuclear destined
segments, or MDSs. The MDSs have become disordered, or scrambled, during evolution. The scrambled structures of the αTP genes
in Oxytricha trifallax and S. mytilus have been compared with the previously published scrambled structure of the αTP gene in O. nova. The scrambled patterns of the αTP gene in the three species are similar but show significant differences. The micronuclear
genes in O. nova and S. mytilus consist of 13 IESs and 14 MDSs, but the gene in O. trifallax is divided into three additional MDSs by the presence of three additional IESs, believed to have been inserted into the O. trifallaxαTP gene after divergence of O. trifallax from the other two species. Corresponding IESs among the three species have shifted along the DNA during evolution, presumably
by a mutational mechanism that changes the short repeat sequences that flank IESs. The IESs also have changed markedly in
length by insertion and/or deletion of nucleotides. Comparison of the putative αTP amino acid sequences in the three species
reveals three conserved and three nonconserved domains. The 5′ nontranslated regions of the gene-sized molecules encoding
αTP contain several conserved segments, and the 3′ nontranscribed trailer contains one conserved segment.
Received: 29 May 1998; in revised form: 3 August 1998 / Accepted: 18 August 1998 相似文献
19.
A gene encoding the trehalose phosphorylase (TreP), which reversibly catalyzes trehalose degradation and synthesis from α-glucose-1-phosphate
(α-Glc-1-P) and glucose, was cloned fromThermoanaerobacter tengcongensis and successfully expressed inEscherichia coli. The overexpressed TreP, with a molecular mass of approximately 90 kDa, was determined by SDS-PAGE. It catalyzes trehalose
synthesis and degradation optimally at 70°C (for 30 min), with the optimum pHs at 6.0 and 7.0, respectively. It is highly
thermostable, with a 77% residual activity after incubation at 50°C for 7 h. Under the optimum reaction conditions, 50 μg
crude enzyme of the TreP is able to catalyze the synthesis of trehalose up to 11.6 mmol/L from 25 mmol/L α-Glc-1-P and 125
mmol/L glucose within 30 min, while only 1.5 mmol/L out of 250 mmol/L trehalose is degraded within the same time period. Dot
blotting revealed that thetreP gene inT. tengcongensis was upregulated in response to salt stress but downregulated when trehalose was supplied. Both results indicate that the
dominant function of theT. tengcongensis TreP is catalyzing trehalose synthesis but not degradation. Thus it might provide a novel route for industrial production
of trehalose. 相似文献
20.
Jian-Hua Chen Xin-Guo Zhang Yu-tao Jiang Lu-Ying Yan Li Tang Yi-Wei Yin Dai-Shuang Cheng Jing Chen Min Wang 《Cancer immunology, immunotherapy : CII》2010,59(9):1335-1345
Thymosin-α1 (Tα1) is indicated for the treatment of certain viral infections, including hepatitis B and C, and cancers, such
as melanoma. In this paper, the fusion genes encoding human serum albumin (HSA) and Tα1 with (rHSA-L-Tα1) and without a linker
peptide (rHSA-Tα1) were constructed and overexpressed in P. pastoris. Through the process of ion interaction chromatography (Q-Sepharose F.F), hydrophobic interaction chromatography (Phenyl
Sepharose HP) and affinity chromatography (Blue Sepharose F.F), the purity of fusion proteins was greater than 97%. In contrast
to the reactivity of normal spleen cells to Con A, the data of in vitro murine spleen lymphocytes proliferation experiment
suggested that spleen cells achieved a higher degree of T cell maturation after rHSA-L-Tα1, rHSA-Tα1 and Tα1 treatments, respectively.
Moreover, rHSA-L-Tα1, rHSA-Tα1 and Tα1 can also antagonize dexamethasone-induced apoptosis of thymocyte sub-populations. In
hydrocortisone-induced immunosuppression mice (in vivo experiments), after subcutaneous injections with two fusion proteins
and Tα1 for seven consecutive days, the net increment of body weight, the spleen index and the thymus index were significantly
improved. Simultaneously, the increase in SOD level and the decrease in MDA level in plasma were observed. The pharmacokinetic
data of rHSA-L-Tα1 and rHSA-Tα1 administered in rats showed an improved pharmacokinetic profile with a conspicuous prolonged
half life. The analysis of bioactivity and pharmacokinetics suggested that fusion proteins rHSA-L-Tα1 and rHSA-Tα1 were new
drug candidates. 相似文献