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1.
Livingstone JR Yoshida I Tarui Y Hirooka K Yamamoto Y Tsutui N Hirasawa E 《Journal of plant research》2002,115(5):393-400
NAD-dependent aminoaldehyde dehydrogenase (AMADH, EC 1.2.1.-) from Avena shoots was purified by DEAE Sephacel, hydroxyapatite, 5′-AMP Sepharose 4B, Mono Q, and TSK-GEL column chromatographies to
homogeneity by the criterion of native PAGE. SDS–PAGE yielded a single band at a molecular mass of 55 kDa. IEF studies showed
a band with a pI value of 5.3. In contrast to AMADHs from other species, the TSK-GEL chromatography showed that Avena AMADH exists as a monomer in the native state. The purified enzyme catalyzed the oxidations of 3-aminopropionaldehyde (APAL),
4-aminobutyraldehyde (ABAL) N-(3-aminopropyl)-4-aminobutyraldehyde (APBAL), and 4-guanidinobutyraldehyde (GBAL), but not of betaine aldehyde or indoleacetaldehyde.
The K
m values for APAL, ABAL, and GBAL were 1.5×10–6, 2.2×10–6, and 1.3×10–5 M, respectively. Although N-terminal amino acid sequence of Avena AMADH could not be determined due to a modification of the amino residue, the sequence of the fragment of AMADH cleaved by
V8 protease showed greater similarity to the barley BADH than to the pea AMADH.
Electronic Publication 相似文献
2.
Suaeda japonica Makino belonging to the family Chenopodiaceae, is a halophyte and grows at the shore of Ariake sea in Japan. This plant presumably
possesses high salt resistant nature, thus, we examined the mechanisms of seed germination under salt stress. The seeds maintained
80% germination rates on the medium containing 0.7 M NaCl. Germination rates varied depending on salt type; the germination
rates under NaCl or KCI exhibited relatively lower values than ones under sodium gluconate or potassium gluconate. This different
responses for salts seemed to be as a result of the presence of Cl− ions. Although very high levels of betaine (compatible solute), were kept in the seedlings grown under no salt stress, the
contents gradually increased as concentration of NaCl increased. Betaine is a factor present in plants that works to alleviate
the effects of excessive soil salts. It is synthesized in leaves from betaine aldehyde, and this process is catabolized by
betaine aldehyde dehydrogenase (BADH). When the seedlings were cultivated on the medium without NaCl, relatively high level
of BADH activity was found. The activity increased 5-fold in the seedlings grown under 0.5 M NaCl stress. Increases in betaine
content and BADH activity were found during seed germination. InS. japonica, the salt stress promoted BADH activity, subsequently endogenous betaine contents were increased, and increased betaine seemed
to secure seed germination under salt stress. 相似文献
3.
Comparative biochemical and immunological studies of the glycine betaine synthesis pathway in diverse families of dicotyledons 总被引:10,自引:0,他引:10
Elizabeth A. Weretilnyk Sebastian Bednarek Kent F. McCue David Rhodes Andrew D. Hanson 《Planta》1989,178(3):342-352
Members of the Chenopodiaceae can accumulate high levels (>100 mol·(g DW)-1) of glycine betaine (betaine) in leaves when salinized. Chenopodiaceae synthesize betaine by a two-step oxidation of choline (cholinebetaine aldehyde betaine), with the second step catalyzed by betaine aldehyde dehydrogenase (BADH, EC 1.2.1.8). High betaine levels have also been reported in leaves of species from several distantly-related families of dicotyledons, raising the question of whether the same betaine-synthesis pathway is used in all cases.Fast atom bombardment mass spectrometry showed that betaine levels of >100 mol·(g DW)-1 are present in Lycium ferocissimum Miers (Solanaceae), Helianthus annuus L. (Asteraceae), Convolvulus arvensis L. (Convolvulaceae), and Amaranthus caudatus L. (Amaranthaceae), that salinization promotes betaine accumulation in these plants, and that they can convert supplied choline to betaine aldehyde and betaine. Nicotiana tabacum L. and Lycopersicon lycopersicum (L.) Karst. ex Farw. (Solanaceae), Lactuca sativa L. (Asteraceae) and Ipomoea purpurea L. (Convolvulaceae) also contained betaine, but at a low level (0.1–0.5 mol·(g DW)-1. Betaine aldehyde dehydrogenase activity assays, immunotitration and immunoblotting demonstrated that the betaine-accumulating species have a BADH enzyme recognized by antibodies raised against BADH from Spinacia oleracea L. (Chenopodiaceae), and that the Mr of the BADH monomer is in all cases close to 63 000. These data indicate that the cholinebetaine aldehydebetaine pathway may have evolved by vertical descent from an early angiosperm ancestor, and might be widespread (albeit not always strongly expressed) among flowering plants. Consistent with these suggestions, Magnolia x soulangiana was found to have a low level of betaine, and to express a protein of Mr 63 000 which cross-reacted with antibodies to BADH from Spinacia oleracea.Abbreviations BADH
Betaine aldehyde dehydrogenase
- DCIMS
desorption chemical ionization mass spectrometry
- FABMS
fast atom bombardment mass spectrometry
- Mr
relative molecular mass
- PAGE
polyacrylamide gel electrophoresis
- SDS
sodium dodecyl sulfate
- TLC
thin-layer chromatography 相似文献
4.
Certain higher plants synthesize and accumulate glycine betaine, a compound with osmoprotectant properties. Biosynthesis of glycine betaine proceeds via the pathway choline betaine aldehyde glycine betaine. Plants such as tobacco (Nicotiana tabacum L.) which do not accumulate glycine betaine lack the enzymes catalyzing both reactions. As a step towards engineering glycine betaine accumulation into a non-accumulator, spinach and sugar beet complementary-DNA sequences encoding the second enzyme of glycine-betaine synthesis (betaine aldehyde dehydrogenase, BADH, EC 1.2.1.8) were expressed in tobacco. Despite the absence of a typical transit peptide, BADH was targeted to the chloroplast in leaves of transgenic plants. Levels of extractable BADH were comparable to those in spinach and sugar beet, and the molecular weight, isoenzyme profile and K
m for betaine aldehyde of the BADH enzymes from transgenic plants were the same as for native spinach or sugar beet BADH. Transgenic plants converted supplied betaine aldehyde to glycine betaine at high rates, demonstrating that they were able to transport betaine aldehyde across both the plasma membrane and the chloroplast envelope. The glycine betaine produced in this way was not further metabolized and reached concentrations similar to those in plants which accumulate glycine betaine naturally. Betaine aldehyde was toxic to non-transformed tobacco tissues whereas transgenic tissues were resistant due to detoxification of betaine aldehyde to glycine betaine. Betaine aldehyded ehydrogenase is therefore of interest as a potential selectable marker, as well as in the metabolic engineering of osmoprotectant biosynthesis.Abbreviations BADH
betaine aldehyde dehydrogenase
- bp
base pairs
- FAB-MS
fast atom bombardment-mass spectrometry
- GAPDH
NADP-linked glyceraldehyde-3-phosphate dehydrogenase
We thank Dr. G. An for the gift of the vector pGA643 and Mr. Sylvain Lebeurier for help in maintaining plants. This work was supported, in part, by grants from the Natural Sciences and Engineering Research Council of Canada, the Rockefeller Foundation, and the U.S. Department of Agriculture, and by gifts from CIBAGEIGY Biotechnology. 相似文献
5.
Toshikazu Chiyonobu Osao Adachi Minoru Ameyama 《Bioscience, biotechnology, and biochemistry》2013,77(9):1743-1744
The significant betaine aldehyde dehydrogenase activity was found in the cells of Pseudomonas aeruginosa A-16. The enzyme was inducibly formed and accumulated in the presence of choline, acetylcholine or betaine in the medium. The enzyme was purified approximately 620-fold with an overall recovery of 2.6% and proved to be homogeneous by ultracentrifugation. The molecular weight of the enzyme was determined as approximately 145,000 by gel filtration method. The enzyme had an isoelectric point around pH 5.1. The enzyme was quite specific for its substrate, betaine aldehyde. Both NADP and NAD functioned as coenzyme. The estimated values of Km at pH 7.4 and 25°C were 3.8 × 10?4 m for betaine aldehyde, 8.9 × 10?5 m for NADP and 2.2 × 10?4 m for NAD. 相似文献
6.
Giovanni Testore Sebastiano Colombatto Francesca Silvagno Stefano Bedino 《The international journal of biochemistry & cell biology》1995,27(11):1201-1210
Oxidative deamination of putrescine, the precursor of polyamines, gives rise to γ-aminobutyraldehyde (ABAL). In this study an aldehyde dehydrogenase, active on ABAL, has been purified to electrophoretic homogeneity from rat liver cytoplasm and its kinetic behaviour investigated. The enzyme is a dimer with a subunit molecular weight of 51,000. It is NAD+-dependent, active only in the presence of sulphhydryl compounds and has a pH optimum in the range 7.3–8.4. Temperatures higher than 28°C promote slow activation and the process is favoured by the presence of at least one substrate. Km for aliphatic aldehydes decreases from 110 μM for ABAL and acetaldehyde to 2–3 μM for capronaldehyde. The highest relative V-values have been observed with ABAL (100) and isobutyraldehyde (64), and the lowest with acetaldehyde (14). Affinity for NAD+ is affected by the aldehyde present at the active site: Km for NAD+ is 70 μM with ABAL, 200 μM with isobutyraldehyde and capronaldehyde, and>800 μM with acetaldehyde. The kinetic behaviour at 37°C is quite complex; according to enzymatic models, NAD+ activates the enzyme (Kact 500 μM) while NADH competes for the regulatory site (Kin 70 μM). In the presence of high NAD+ concentrations (4 mM), ABAL promotes further activation by binding to a low-affinity regulatory site (Kact 10 mM). The data show that the enzyme is probably an E3 aldehyde dehydrogenase, and suggest that it can effectively metabolize aldehydes arising from biogenic amines. 相似文献
7.
Betaine content in leaves of fifteen plant species was determined. The results showed higher betaine levels in those salt-, drought-, and chilling-resistant species. Betaine aldehyde dehydrogenase (BADH, EC 1.2.1.8 ) was isolated and partially purified from spinach leaves. Some properties of this enzyme were studied. BADH was precipitated by 60% saturation of (NH4)2SO4. Its activity was not detected in 70% saturation of (NH4)2SO4. BADH has two isoenzymes. The activity of BADH was quite stable below –80℃. It was inhibited by 0.125–1.0 mol/L NaG1 or KC1 but not by Mn2+ and Mo6+, and slightly increased by Mg2+. 相似文献
8.
Csar Muoz‐Bacasehua Jesús A. Rosas‐Rodríguez Aldo A. Arvizu‐Flores Aurora Stephens‐Camacho Jos G. Soanez‐Organis Ciria G. Figueroa‐Soto Elisa M. Valenzuela‐Soto 《Journal of molecular recognition : JMR》2020,33(10)
Betaine aldehyde dehydrogenase (BADH EC 1.2.1.8) catalyzes the irreversible oxidation of betaine aldehyde to glycine betaine using NAD+ as a coenzyme. Porcine kidney BADH (pkBADH) follows a bi‐bi ordered mechanism in which NAD+ binds to the enzyme before the aldehyde. Previous studies showed that NAD+ induces complex and unusual conformational changes on pkBADH and that potassium is required to maintain its quaternary structure. The aim of this work was to analyze the structural changes in pkBADH caused by NAD+ binding and the role played by potassium in those changes. The pkBADH cDNA was cloned and overexpressed in Escherichia coli, and the protein was purified by affinity chromatography using a chitin matrix. The pkBADH/NAD+ interaction was analyzed by circular dichroism (CD) and by isothermal titration calorimetry (ITC) by titrating the enzyme with NAD+. The cDNA has an open reading frame of 1485 bp and encodes a protein of 494 amino acids, with a predicted molecular mass of 53.9 kDa. CD data showed that the binding of NAD+ to the enzyme caused changes in its secondary structure, whereas the presence of K+ helps maintain its α‐helix content. K+ increased the thermal stability of the pkBADH‐NAD+ complex by 5.3°C. ITC data showed that NAD+ binding occurs with different association constants for each active site between 37.5 and 8.6 μM. All the results support previous data in which the enzyme incubation with NAD+ provoked changes in reactivity, which is an indication of slow conformational rearrangements of the active site. 相似文献
9.
Several natural products were tested to control the differentiation of cultured human mesenchymal stem cell into adipocyte.
The extent of the inhibitory effect on the conversion of adipose was measured using Oil red O staining, which stains accumulated
lipid droplets in the cytoplasm of adipocyte. Of the various natural product extracts, the adipocyte differentiation was inhibited
by an extract fromFolium mori in the concentration range 1×10−4∼5×10−2 g/mL. These results suggest thatFolium mori has an inhibitory activity toward the adipose conversion, which is a major cause of obesity. 相似文献
10.
Kemel Jellouli Ali Bougatef Laila Manni Rym Agrebi Rayda Siala Islem Younes Moncef Nasri 《Journal of industrial microbiology & biotechnology》2009,36(7):939-948
A protease-producing bacterium was isolated from an alkaline wastewater of the soap industry and identified as Vibrio metschnikovii J1 on the basis of the 16S rRNA gene sequencing and biochemical properties. The strain was found to over-produce proteases
when it was grown at 30°C in media containing casein as carbon source (14,000 U ml−1). J1 enzyme, the major protease produced by V. metschnikovii J1, was purified by a three-step procedure, with a 2.1-fold increase in specific activity and 33.3% recovery. The molecular
weight of the purified protease was estimated to be 30 kDa by SDS-PAGE and gel filtration. The N-terminal amino acid sequence
of the first 20 amino acids of the purified J1 protease was AQQTPYGIRMVQADQLSDVY. The enzyme was highly active over a wide
range of pH from 9.0 to 12.0, with an optimum at pH 11.0. The optimum temperature for the purified enzyme was 60°C. The activity
of the enzyme was totally lost in the presence of PMSF, suggesting that the purified enzyme is a serine protease. The kinetic
constants K
m and K
cat of the purified enzyme using N-succinyl-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide were 0.158 mM and 1.14 × 105 min−1, respectively. The catalytic efficiency (K
cat
/K
m) was 7.23 × 108 min−1 M−1. The enzyme showed extreme stability toward non-ionic surfactants and oxidizing agents. In addition, it showed high stability
and compatibility with some commercial liquid and solid detergents. The aprJ1 gene, which encodes the alkaline protease from V. metschnikovii J1, was isolated, and its DNA sequence was determined. The deduced amino acid sequence of the preproenzyme differs from that
of V. metschnikovii RH530 detergent-stable protease by 12 amino acids, 7 located in the propeptide and 5 in the mature enzyme. 相似文献
11.
Longstreth David J. Burow Gloria B. Yu Gang 《Plant Cell, Tissue and Organ Culture》2004,78(3):225-230
Cell recovery from osmotic stress was studied in suspension cell cultures from Alternanthera philoxeroides [Mart.] Griseb. Changes in different classes of cellular solutes were measured after cells were transferred from 0 to 200
mM NaCl (high salt) to obtain an integrated picture of the solute pools involved in osmotic adjustment. By 2 h, cellular [Na+] and [Cl−] had increased several-fold, potentially accounting for the osmotic adjustment that produced a rapid recovery of cell turgor.
There was a four-fold increase in the concentration of quaternary ammonium compounds (QAC) by 12 h and a slower increase for
several days afterward. Betaine aldehyde dehydrogenase (BADH) is required for synthesis of glycine betaine, a QAC produced
by a range of organisms in response to osmotic stress. Western-blot analysis for BADH suggested that glycine betaine was a
significant component of the QAC solutes. The amount of BADH was generally similar at different sampling times for control
and high salt cells, unlike previous reports of stimulation by osmotic stress in intact plants of some species. Between 3
and 7 days after cell transfer to high salt, other organic solutes increased in concentration and [Na+] and [Cl−] decreased. In A. philoxeroides, high [Na+] and [Cl−] produce rapid osmotic adjustment but organic solutes apparently replace these potentially harmful inorganic ions after the
recovery of turgor. 相似文献
12.
Quaratino D Federici F Petruccioli M Fenice M D'Annibale A 《Antonie van Leeuwenhoek》2007,91(1):57-69
Extracellular laccase from Panus tigrinus CBS 577.79 was produced in a bubble-column reactor using glucose-containing medium supplemented with 2,5-xylidine under conditions of nitrogen sufficiency. The main laccase isoenzyme was purified to apparent homogeneity by ultra-filtration, anion-exchange chromatography and gel filtration that led to a purified enzyme with a specific activity of 317 IU (mg protein)−1 and a final yield of 66%. Laccase was found to be a monomeric protein with a molecular mass of 69.1 kDa, pI of 3.15 and 6.9% N-glycosylation of the high mannose type. Temperature and pH optima were 55°C and 3.75 (2,6-dimethoxyphenol as substrate). At 50 and 60°C, the enzyme half-lives were 281 and 25 min, respectively. The P. tigrinus laccase oxidized a wide range of both naturally occurring and synthetic aromatic compounds: the highest catalytic efficiencies were for 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonic) acid and 2,6-dimethoxyphenol (5.99 × 106 and 3.07 × 106 M−1 s−1, respectively). Catalytic rate constants for typical N–OH redox mediators, such as 1-hydroxybenzotriazole (2.6 s−1), violuric acid (8.4 s−1) and 2,2,6,6-tetramethylpiperidin-N-oxide radical (7.8 s−1), were found to be higher than those reported for other high redox potential fungal laccases. 相似文献
13.
Li-Ping Yan Cui-Lan Liu Hui-Min Liang Xiu-Hong Mao Fei Wang Cai-Hong Pang Jing Shu Yang Xia 《Plant Cell, Tissue and Organ Culture》2012,108(2):191-199
Glycinebetaine is an important quaternary ammonium compound generated in response to salt and other osmotic stresses in many
organisms. Its synthesis requires the catalysis of betaine aldehyde dehydrogenase encoded by a Betaine Aldehyde Dehydrogenase
(BADH) gene that converts betaine aldehyde into glycinebetaine in some halotolerant plants. In this study, a BADH gene was over expressed in transgenic alfalfa (Medicago sativa L) plants using Agrobacterium-mediated transformation. Transgenic alfalfa plants grown under 9‰ NaCl grew well; while non-transgenic
control plants turned yellowish in color, wilted, and eventually died. Polymerase chain reaction (PCR) and Northern blot hybridization
analyses demonstrated that the BADH gene was transferred into the T2 generation and segregated in a Mendelian fashion. Transgenic alfalfa plants expressing BADH showed significantly higher BADH enzyme activity and betaine contents when grown under 6‰ NaCl. Moreover, proline content
in T2 lines were higher while electrolyte leakage and malonaldehyde content were lower in T2 lines compared with non-transgenic
plants. These findings indicated that transgenic plants expressing BADH transgene exhibited higher salt tolerance than non-transgenic plants. 相似文献
14.
J. K. Sharma M. Yadav N. P. Singh K. D. S. Yadav 《Applied Biochemistry and Microbiology》2011,47(5):532-537
Extracellular secretion of lignin peroxidase from Pycnoporus sanguineus MTCC-137 in the liquid culture growth medium amended with lignin containing natural sources has been shown. The maximum secretion
of lignin peroxidase has been found in the presence of saw dust. The enzyme has been purified to homogeneity from the culture
filtrate of the fungus using ultrafiltration and anion exchange chromatography on DEAE-cellulose. The purified lignin peroxidase
gave a single protein band in sodium dodecylsulphate polyacrylamide gel electrophoresis corresponding to the molecular mass
40 kDa. The K
m, k
cat and k
cat/K
m values of the enzyme using veratryl alcohol and H2O2 as the substrate were 61 M, 2.13 s−1, 3.5 × 104 M−1s−1 and 71 M, 2.13 s−1, 3.0 × 104 M−1 s−1 respectively at the optimum pH of 2.5. The temperature optimum of the enzyme was 25°C. 相似文献
15.
Guosheng Shao Mingxue Chen Weixia Wang Guoping Zhang 《Journal of Plant Growth Regulation》2008,27(3):205-210
The influence of betaine aldehyde dehydrogenase (BADH) and salinity pretreatment on oxidative stress under cadmium (Cd) toxicity
was investigated in rice cv. Xiushui 11 and its BADH-transgenic line Bxiushui 11. The results showed that plants previously treated with 4.25 and 8.5 mM NaCl, respectively, for
5 days each had higher Cd concentrations in both roots and shoots of the two rice genotypes compared with the controls. Malondialdehyde
(MDA) content in both leaves and roots was increased by salinity pretreatment and was significantly lower in the salinity-pretreatment
plants than in the controls when the plants were consequently exposed to Cd stress. Salinity pretreatment also increased proline
content and the activities of superoxide dismutase (SOD) and peroxidase (POD) in both leaves and roots. It can be assumed
that salinity pretreatment enhances the defensive ability of plants against oxidative stress through increasing activities
of antioxidative enzymes. The BADH-transgenic line (Bxiushui 11) had lower Cd and MDA content, higher SOD and POD activities, and higher proline content than
its wild type (Xiushui 11). The current results suggest that betaine, a product of BADH expression, improves the tolerance of rice plants to Cd stress through increasing the activities of antioxidative enzymes
and osmoprotectant content. 相似文献
16.
Production of the compatible solute glycine betaine from its precursors choline or glycine betaine aldehyde confers a considerable
level of tolerance against high osmolarity stress to the soil bacterium Bacillus subtilis. The glycine betaine aldehyde dehydrogenase GbsA is an integral part of the osmoregulatory glycine betaine synthesis pathway.
We strongly overproduced this enzyme in an Escherichia coli strain that expressed a plasmid-encoded gbsA gene under T7φ10 control. The recombinant GbsA protein was purified 23-fold to apparent homogeneity by fractionated ammonium
sulfate precipitation, ion-exchange chromatography on Q-Sepharose, and subsequent hydrophobic interaction chromatography on
phenyl-Sepharose. Molecular sieving through Superose 12 and sedimentation centrifugation through a glycerol gradient suggested
that the native enzyme is a homodimer with 53.7-kDa subunits. The enzyme was specific for glycine betaine aldehyde and could
use both NAD+ and NADP+ as cofactors, but NAD+ was strongly preferred. A kinetic analysis of the GbsA-mediated oxidation of glycine betaine aldehyde to glycine betaine
revealed K
m values of 125 μM and 143 μM for its substrates glycine betaine aldehyde and NAD+, respectively. Low concentrations of salts stimulated the GbsA activity, and the enzyme was highly tolerant of high ionic
conditions. Even in the presence of 2.4 M KCl, 88% of the initial enzymatic activity was maintained. B. subtilis synthesizes high levels of proline when grown at high osmolarity, and the presence of this amino acid strongly stimulated
the GbsA activity in vitro. The enzyme was stimulated by moderate concentrations of glycine betaine, and its activity was
highly tolerant against molar concentrations of this osmolyte. The high salt tolerance and its resistance to its own reaction
product are essential features of the GbsA enzyme and ensure that B. subtilis can produce high levels of the compatible solute glycine betaine under conditions of high osmolarity stress.
Received: 2 May 1997 / Accepted: 2 July 1997 相似文献
17.
Wongpanya R Boonyalai N Thammachuchourat N Horata N Arikit S Myint KM Vanavichit A Choowongkomon K 《The protein journal》2011,30(8):529-538
Betaine aldehyde dehydrogenase 2 (BADH2) is believed to be involved in the accumulation of 2-acetyl-1-pyrroline (2AP), one
of the major aromatic compounds in fragrant rice. The enzyme can oxidize ω-aminoaldehydes to the corresponding ω-amino acids.
This study was carried out to investigate the function of wild-type BADHs and four BADH2 mutants: BADH2_Y420, containing a
Y420 insertion similar to BADH2.8 in Myanmar fragrance rice, BADH2_C294A, BADH2_E260A and BADH2_N162A, consisting of a single
catalytic-residue mutation. Our results showed that the BADH2_Y420 mutant exhibited less catalytic efficiency towards γ-aminobutyraldehyde
but greater efficiency towards betaine aldehyde than wild-type. We hypothesized that this point mutation may account for the
accumulation of γ-aminobutyraldehyde/Δ1-pyrroline prior to conversion to 2AP, generating fragrance in Myanmar rice. In addition, the three catalytic-residue mutants
confirmed that residues C294, E260 and N162 were involved in the catalytic activity of BADH2 similar to those of other BADHs. 相似文献
18.
Julio C. Davila Chandrasekara G. Reddy Patrick J. Davis Daniel Acosta 《In vitro cellular & developmental biology. Plant》1990,26(5):515-524
Summary The present study was undertaken to assess and compare the toxic effects of papaverine hydrochloride and its metabolites.
Primary cell cultures of rat hepatocytes were treated with papavarine (papaver), 3′-O-desmethyl (3′-OH), 4′-O-desmethyl (4′-OH),
and 6-O-desmethyl (6-OH) papaverine at 1×10−5, 1×10−4, and 1×10−3
M for 4,8, 12, and 24-h periods. Cell injury was determined by: a) cell viability using the trypan blue exclusion test; b)
cytosolic enzyme leakage of lactate dehydrogenase and aspartate aminotransferase; c) morphologic alterations; and d) lactate:
pyruvate (L:P) ratios. Cell cultures showed concentration-and time-dependent responses. For example, a decrease in cell viability
and an increase in enzyme leakage were observed after cell treatment with 1×10−4 and 1×10−3
M papaver for 8 h; 1×10−3
M 6-OH papaverine for 8 h and 1×10−4
M for 24 h; and 1×10−3
M 4′-OH papaverine for 24 h (P<0.05). Furthermore, changes in morphology correlated to cell viability and enzyme release in those cultures treated with
papaver, 4′-OH and 6-OH papaverine. Some of these changes included size deformation, cell detachment from the dishes, and
cell necrosis. On the other hand, an increase in L:P ratios (P<0.05) was detected with papaver as early as 8 h with 1×10−4 and 1×10−3
M and 12 h with 1×10−5
M; 6-OH showed an increase, in L:P ratios at 8 h with 1×10−3
M and 12 h with 1×10−4
M; these changes were evident with 4′-OH at 12 h with 1×10−3
M. In contrast, cells treated with 3′-OH papaverine did not show significant damage with any time period and concentration
used in this study. The results of this study indicate that papaverine-derived metabolites are less cytotoxic than its parent
compound, papaver. The toxicity was ranked as follows: papaver>6-OH>4′-OH>−3′-OH.
This work was supported in part by grant ES04200-02 from the National Institute of Environmental Health Sciences, Bethesda,
MD.
Presented in part at the fall ASPET meeting in Salt Lake City, August, 1989. Daniel Acosta is a Burroughs Wellcome Scholar
in Toxicology. 相似文献
19.
Madanala R Gupta V Deeba F Upadhyay SK Pandey V Singh PK Tuli R 《Biotechnology letters》2011,33(10):2057-2063
A gene from Withania somnifera (winter cherry), encoding a highly stable chloroplastic Cu/Zn superoxide dismutase (SOD), was cloned and expressed in Escherichia coli. The recombinant enzyme (specific activity of ~4,200 U mg−1) was purified and characterized. It retained ~90 and ~70% residual activities after 1 h at 80 and 95°C, respectively. At
95°C, thermal inactivation rate constant (K
d) of the enzyme was 2.46 × 10−3 min−1 and half-life of heat inactivation was 4.68 h. The enzyme was stable against a broad pH range (2.5–11.0). It also showed
a high degree of resistance to detergent, ethanol and protease digestion. This recombinant Cu/Zn SOD could therefore have
useful applications. 相似文献
20.
Diamine oxidase was purified sixty-fold from millet shoots. The partially purified enzyme of 150 kDa oxidized 1, 3-diaminopropane
(1, 3-DAP) to 3-aminopropionaldehyde. The Km values were 9.1×10−5M for 1, 3-DAP and 6.3×10−4M for putrescine. Extracts of shoots of prosomillet, maize and barley also contained an activity that oxidized 1, 3-DAP. 相似文献