共查询到20条相似文献,搜索用时 78 毫秒
1.
Tony Márcio Silva Fausto Bruno Dos Reis Almeida André Ricardo de Lima Damásio Alexandre Maller Michele Michelin João Atílio Jorge Ebert Seixas Hanna Maria Cristina Roque-Barreira Héctor F. Terenzi Maria de Lourdes Teixeira de Moraes Polizeli 《Biotechnology letters》2010,32(10):1449-1455
Treatment of Aspergillus niveus with 30 μg tunicamycin/ml did not interfere with α-glucosidase production, secretion, or its catalytic properties. Fully-
and under-glycosylated forms of the enzyme had similar molecular masses, ~56 kDa. Moreover, the absence of N-glycans did not affect either pH optimum (6.0) or temperature optimum (65°C). The Km and Vmax values of under- and fully-glycosylated forms of α-glucosidase were similar when assessed for hydrolysis of starch (~0.6 mg/ml,
~350 μmol glucose per min per ml), maltose (~0.54 μmol, ~330 μmol glucose per min per ml) and p-nitrophenyl-α-d-glucopyranoside (~0.54 μmol, ~8.28 μmol p-nitrophenol per min per ml). However, the under-glycosylated form was sensitive to high temperatures probably because, in
addition to stabilizing the protein conformation, glycosylation may also prevent unfolded or partially folded proteins from
aggregating. Binding assays clearly showed that the under-glycosylated protein did not bind to concanavalin A but has conserve
its jacalin-binding property, suggesting that only O-glycans might be intact on the tunicamycin treated form of the enzyme. 相似文献
2.
Villalonga R Fujii A Shinohara H Asano Y Cao R Tachibana S Ortiz P 《Biotechnology letters》2007,29(3):447-452
A supramolecular approach was used for adsorbing a monolayer of adamantane-modified phenylalanine dehydrogenase on β-cyclodextrin-coated Au electrodes. The enzyme electrode (poised at +200 mV vs. Ag/AgCl) showed a linear amperometric response
up to 3 mM l-phenylalanine (l-Phe) with a lower detection limit of 15 μM. The reversible nature of this immobilization approach was confirmed. 相似文献
3.
In addition to the general amino acid transport system (GAP) ofS. cerevisiae
l-tryptophan is transported by another system with approximately 25% capacity of GAP, with aK
T of 0.41±0.08 mmol/L and with a similar specificity as GAP (lower inhibition by Met, Pro, Ser, Thr and 2-aminoisobutyric acid;
greater inhibition by Glu and His). The pH optimum of this system is at 5.0–5.5, activation energy above the transition point
(20°C) was 20 kJ/mol, below the transition point 55 kJ/mol. The transport by this system was virtually unidirectional, efflux
amounting to at most 10% into a tryptophan-free medium. The transport itself was blocked by 2,4-dinitrophenol, antimycin A
and uranyl nitrate. The system was synthesized de novo during preincubation with glucose=fructose>trehalose >ethanol within
30 min, and was degraded with a half-time of 15 min in the absence of further synthesis. The accumulation ratios ofl-tryptophan ingap1 mutants were concentration-dependent (200∶1 at 1 μmoll-Trp/L, 4∶1 at 2.5 mmoll-Trp/L) and decreased with increasing suspension density from 200∶1 to 5∶1 (for 10 μmoll-Trp/L). The involvement of hydrogen ions in the uptake was clearly demonstrated by the effect of D2O even if it could not be established by either shifts of pHout or membrane depolarization. 相似文献
4.
M. Torras-Llort R. Ferrer J.F. Soriano-García M. Moretó 《The Journal of membrane biology》1996,152(3):183-193
The properties of l-lysine transport in chicken jejunum have been studied in brush border membrane vesicles isolated from 6-wk-old birds. l-lysine uptake was found to occur within an osmotically active space with significant binding to the membrane. The vesicles
can accumulate l-lysine against a concentration gradient, by a membrane potential-sensitive mechanism. The kinetics of l-lysine transport were described by two saturable processes: first, a high affinity-transport system (K
mA= 2.4 ± 0.7 μmol/L) which recognizes cationic and also neutral amino acids with similar affinity in the presence or absence
of Na+ (l-methionine inhibition constant KiA, NaSCN = 21.0 ± 8.7 μmol/L and KSCN = 55.0 ± 8.4 μmol/L); second, a low-affinity transport mechanism (KmB= 164.0 ± 13.0 μmol/L) which also recognizes neutral amino acids. This latter system shows a higher affinity in the presence
of Na+ (KiB for l-methionine, NaSCN = 1.7 ± 0.3 and KSCN = 3.4 ± 0.9 mmol/L). l-lysine influx was significantly reduced with N-ethylmaleimide (0.5 mmol/L) treatment. Accelerative exchange of extravesicular labeled l-lysine was demonstrated in vesicles preloaded with 1 mmol/L l-lysine, l-arginine or l-methionine. Results support the view that l-lysine is transported in the chicken jejunum by two transport systems, A and B, with properties similar to those described
for systems b
0,+ and y+, respectively.
Received: 14 August 1995/Revised: 2 April 1996 相似文献
5.
Effect of Abscisic Acid on Banana Fruit Ripening in Relation to the Role of Ethylene 总被引:7,自引:0,他引:7
Abstract
The role of abscisic acid (ABA) in banana fruit ripening was examined with the ethylene binding inhibitor, 1-methylcyclopropene
(1-MCP). ABA (0, 10−5, 10−4, or 10−3 mol/L) was applied by vacuum infiltration into fruit. 1-MCP (1 μL/L) was applied by injecting a measured volume of stock
gas into sealed glass jars containing fruit. Fruit ripening, as judged by ethylene evolution and respiration associated with
color change and softening, was accelerated by 10−4 or 10−3 mol/L ABA. ABA at 10−5 mol/L had no effect. The acceleration of ripening by ABA was greater at 10−3 mol/L than at 10−4 mol/L. ABA-induced acceleration of banana fruit ripening was not observed in 1-MCP treated fruit, especially when ABA was
applied after exposure to 1-MCP. Thus, ABA's promotion of ripening in intact banana fruit is at least partially mediated by
ethylene. Exposure of ABA-treated fruit to 0.1 μL/L ethylene for 24 h resulted in increased ethylene production and respiration,
and associated skin color change and fruit softening. Control fruit (no ABA) was unresponsive to similar ethylene treatments.
The data suggest that ABA facilitates initiation and progress in the sequence of ethylene-mediated ripening events, possibly
by enhancing the sensitivity to ethylene.
Received 29 January 1999; accepted 16 January 2000 相似文献
6.
Julius Brtko Peter Filipčík Soňa Hudecová Anastázia Brtková Janette Bransová 《Biological trace element research》1998,62(1-2):43-50
The present study was undertaken to investigate the effects of selenite (SeIV) and selenate (SeVI) on the all-trans retinoic acid (RA)-nuclear retinoic acid receptor (RAR) complex formation in rat liver. We also present
the data on the in vitro effects of SeIV on the RARα and the type I iodothyronine 5′-deiodinase gene expression in the GH4C1 rat pituitary tumor cells. SeIV at 1.0 μmol/L was found to reduce (p<0.05) the RA specific binding to RAR in rat liver. Dithiothreitol (DTT), a protective agent for sulfhydryl groups, was found
to be slightly effective in protecting the RAR binding properties when affected by SeIV. SeVI at 0.1 μmol/L reduced (p<0.05) the RA specific binding to RAR in liver, as well. Seleno-l-methionine (Se-II) when compared tol-methionine did not exert any inhibitory effect on the formation of the RA-RAR complex. SeIV (up to 2.5 μmol/L) has no inhibitory effect on GH4C1 cell proliferation as well as the prolactin secretion. SeIV at 1.0 μmol/L significantly decreases the rate of mRNA synthesis and/or degradation of the α form of the RAR and causes the
enhancement of the type I iodothyronine 5′-deiodinase gene expression in GH4C1 cells.
The results based on in vitro experiments suggest that inorganic selenium may affect the RA specific binding to their cognate
receptor molecules, and it may reduce expression of the gene encoding the RARα, with the cell vitality and the cell growth
remaining unchanged. 相似文献
7.
An extracellular endo-d-arabinase enzyme produced by the bacterial strain of Cellulomonas was purified 77.1-fold with 0.20% recovery for protein by DEAE Sepharose anion exchange, Sephacryl S-300 gel filtration and
blue Sepharose affinity chromatography, and designated as CEDAase. The apparent molecular mass of CEDAase was 45 kDa determined
by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. CEDAase is an endoenzyme for arabinogalactan with the main and
specific product of hexa-arabinofuranoside. It reacts optimally with its substrate, arabinogalactan, at approximately pH 8.0
and at 40 °C. CEDAase shows stability in the pH range of 6.0–9.0 and at the temperature below 50 °C. The Km measured for the CEDAase was 55.6 μM, with an apparent Vmax of 0.083 μmol/min. To our knowledge, for the first time, the current work obtains an extracellular Cellulomonas endo-d-arabinase enzyme that might be potentially served as a tool enzyme for hydrolyzing specific cell wall such as Mycobacterium cell. It is purified as an important potential initial material basis for mass spectrometric sequencing and chemical gene
synthesis. It may make it possible to clone and express this valuable endo-d-arabinase and make it available to the mycobacteria scientific community. 相似文献
8.
Law RO 《Neurochemical research》2005,30(12):1465-1470
Cell volumes (equilibrium non-inulin spaces) have been measured in slices of rat cerebral cortex incubated in the presence
of uraemic guanidino compounds. Of 5 guanidino compounds tested, all but one caused significant cell swelling. This was most
pronounced for guanidinosuccinic acid (GSA, 40 μmol/l)(+22%) and guanidine hydrochloride (G, 3 μmol/l)(+13%). Swelling was
reduced by taurine in a dose-dependent manner, being completely abolished at 20 mmol/l. Swelling was also abolished by the
antioxidants ascorbic acid (0.4 mmol/l) and butylated hydroxytoluene (0.5 mmol/l), the free radical scavenger N-acetyl-l-cysteine (10 mmol/l) and the lipid peroxidase inhibitor desmethyl tirilazad (100 μmol/l). The remission of swelling by 20 mmol/l
taurine was reduced by 50% by the taurine transport inhibitor guanidinoethylsulphonate (GES, 1 mmol/l). This figure was not
significantly altered when the concentration of GES was increased to 10 mmol/l. It was also reduced by 45% by the GABAa receptor antagonist bicuculline (100 μmol/l). It was completely abolished when both GES and bicuculline were present. It
is suggested that guanidino compounds result in cells undergoing oxidative-nitrosative stress, and that taurine protects against
the resultant cell swelling by 2 mechanisms One (intracellular) requires taurine transport and depends on its role as an antioxidant,
with lipid peroxidation being probably a significant factor. The other (extracellular) is associated with activation of GABAa receptors.
Special issue dedicated to Simo S. Oja 相似文献
9.
Riboflavin, suggested to be a radiosensitizer, was studied in murine thymocytes and human hepatoma L02 cell line in vitro with MTT method and fluorescence microscopy. When the murine thymocytes treated with 5–400 μmol/L riboflavin were irradiated
by 5 Gy 60Co γ ionizing radiation, the low concentration groups, i.e. treated with 5–50 μmol/L riboflavin, showed a different surviving
fractions-time relating correlation compared with the high concentration groups, i.e. treated with 100–400 μmol/L riboflavin.
The former had a high survival level at the end of irradiation, but which, after 4-h incubation, decreased rapidly to a low
level. On the contrary, the high concentration groups showed a low survival level at the end of irradiation, and a poor correlation
was found between the surviving fraction and the incubation time, after 4 h a little difference was observed. The results
of fluorescence microscopy indicated that under low concentration conditions, the riboflavin localized mainly in nucleus (both
perinuclear area and inside of nuclear membrane), while under high concentration conditions, intensive riboflavin also localized
around cytoplasmic membranes. Thus we can conclude: the riboflavin had radiosensitivity effect on DNA under low concentration
conditions, and enhanced the damage to cytoplasmic membrane under high concentration conditions. Also the most effective concentration
of riboflavin can be evaluated to be approximate 100 μmol/L. 相似文献
10.
5′-O-β-d-Galactosyl-floxuridine, a potential novel prodrug, was synthesized with a yield of 75% through β-galactosidase-catalyzed transgalactosylation. This enzyme displayed absolute regioselectivity toward the 5′-position of floxuridine.
For the reaction, the optimal conditions were pH 6.5 at 45°C for 60 h with floxuridine to o-nitrophenyl-β-d-galactoside at 2:1 (mol/mol). Under these conditions, the initial reaction rate and the maximum yield were 0.28 mM h−1 and 75%, respectively. 相似文献
11.
Y. W. Kim R. Newton J. Frampton K.-H. Han 《In vitro cellular & developmental biology. Plant》2009,45(4):400-406
Embryogenic suspensor mass (ESM) was established from immature seeds of Fraser fir. The initiation frequency of ESM was dependent
on genotype, collection time, medium, and plant growth regulators (PGR) used. The ESM initiation potential was higher with
seeds collected in late June (clone 16-273, 4.7%) or early July (clone 16-45, 2.2%) and decreased as the zygotic embryos matured.
Excised proembryo stage of zygotic embryos was most appropriate to initiation of ESM. Most of the ESM arose from the seeds
that were at the proembryo stage. From the four different culture media we compared, seven ESM lines were obtained: two lines
from Murashige and Skoog (MS) medium with 4.4 μM benzyladenine (BA), one from Schenk and Hildebrandt (SH) medium with 4.5 μM
thidiazuron (TDZ), and four from SH with 4.4 μM 6-benzyladenine. However, only one ESM line from clone 16-273 (June 24, SH+TDZ)
could be proliferated in subsequent culture. Different concentrations of l-glutamine and casein hydrolysate (CH) in the medium were also compared for their effect on ESM proliferation. The highest
proliferation rate (1.16-fold) was obtained from SH medium supplemented with 250 mg/L CH and 3.42 mM l-glutamine. In contrast, the lowest rate was noted when 1,000 mg/L CH plus 3.42 mM l-glutamine (0.17-fold) was added to the medium. As for somatic embryo maturation, the highest number of mature precotyledonary
(100.1/g−1 FW ESM) or cotyledonary (64.3/g−1 FW ESM) somatic embryos was obtained on a medium containing 20 or 80 μM abscisic acid, 10% polyethyleneglycol, 4% maltose,
and 0.3% gellan gum. For germination of the somatic embryos, the cotyledonary somatic embryos derived from maturation medium
were transferred on half-strength Litvay medium containing 0.3% gellan gum. The somatic plantlets were recovered from the
germination medium and transferred to soils. 相似文献
12.
Olga Molchan Svetlana Romashko Vladimir Yurin 《Plant Cell, Tissue and Organ Culture》2012,108(3):535-539
l-tryptophan decarboxylase (TDC, EC 4.1.1.28) catalyses the formation of tryptamine from tryptophan, and therefore it plays
a role in terpenoid indole alkaloids biosynthesis. In this study, TDC activity and tryptamine accumulation were monitored
in callus cultures of important medicinal plant Vinca minor L. Callus cultures, established from leaf tissues, were incubated on Murashige and Skoog (MS) medium supplemented with 4.4 μM
kinetin and different concentrations (0.44, 1.1, 2.2, 4.4 and 6.6 μM) of naphthaleneacetic acid (NAA), and grown either in
the dark or under 16 h photoperiod. When the basal enzyme activity of TDC was determined in these cultures, it was 0.5–0.7 nmol
tryptamine mg−1 prot. min−1. Moreover, this activity remained linear over time and over protein concentrations, and with optimum pH levels between 6.5
and 7.5, and an optimum temperature of 35°C. The Michaelis–Menten constant (Km) for l-tryptophan was 1.3 mM. TDC cofactor, pyridoxal-5′-phosphate (1 mM), increased the enzyme activity. During later stages of
callus culture growth cycle, an increase in TDC activity was observed, and this activity depended on culture conditions and
age of callus cultures. In addition, TDC activity and tryptamine accumulation in callus cultures were strongly enhanced by
light treatment. 相似文献
13.
Mahdi Mohammadian Mehrnoosh Fathi-Roudsari Nasrin Mollania Arastoo Badoei-Dalfard Khosro Khajeh 《Journal of industrial microbiology & biotechnology》2010,37(8):863-869
Laccases (benzenediol oxygen oxidoreductase; EC 1.10.3.2) have many biotechnological applications because of their oxidation
ability towards a wide range of phenolic compounds. Within recent years, researchers have been highly interested in the identification
and characterization of laccases from bacterial sources. In this study, we have isolated and cloned a gene encoding laccase
(CotA) from Bacillus sp. HR03 and then expressed it under microaerobic conditions and decreased temperature in order to obtain high amounts of
soluble protein. The laccase was purified and its biochemical properties were investigated using three common laccase substrates,
2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), syringaldazine (SGZ) and 2,6-dimethoxyphenol (2,6-DMP). K
M and k
cat were calculated 535 μM and 127 s−1 for ABTS, 53 μM and 3 s−1 for 2, 6-DMP and 5 μM and 20 s−1 for SGZ when the whole reactions were carried out at room temperature. Laccase activity was also studied when the enzyme
was preincubated at 70 and 80°C. With SGZ as the substrate, the activity was increased three-fold after 50 min preincubation
at 70°C and 2.4-fold after 10 min preincubation at 80°C. Preincubation of the enzyme in 70°C for 30 min raised the activity
four-fold with ABTS as the substrate. Also, l-dopa was used as a substrate. The enzyme was able to oxidize l-dopa with the K
M and k
cat of 1,493 μM and 194 s−1, respectively. 相似文献
14.
Cell-free extracts (CFEs) of chromium-resistant bacterium Bacillus sphaericus AND 303 isolated from serpentine soil of Andaman, India reduced Cr(VI) in in vitro condition, and the reductase activity was solely localized in the soluble cell-fractions (S12, S32, and S150). The enzyme was constitutive as the CFEs from cells grown in Cr(VI)-free and Cr(VI)-containing media reduced a more or less
equal amount of Cr(VI). Optimum Cr(VI) reductase activity was obtained at an enzyme (S150) concentration equivalent to 4.56 mg protein/mL, 300 μM Cr(VI) and pH 6.0 after 30 min incubation at 30°C. The enzyme was heat labile; 80% of its activity was lost when exposed
at 70°C for 15 min. Kinetics of Cr(VI) reductase activity fit well with the linearized Lineweaver-Burk plot and showed a Vmax of 1.432 μmol Cr(VI)/mg protein/min and Km of 158.12 μM Cr(VI). The presence of additional electron donors accelerated Cr(VI) reductase activity of CFE, and an increase of 28% activity
over control was recorded with 1.0 μM NADH. Heavy metal ions such as Ni(II), Cu(II), and Cd(II) were strong inhibitors of Cr(VI) reductase unlike that of 100 μM Co(II), which retained 93% activity over control. 相似文献
15.
Ziguo Zhao Fengjuan Zhao Jianting Yao Jingmei Lu Put. O. AngJr. Delin Duan 《Journal of applied phycology》2008,20(5):925-931
Morphology and culture studies on germlings of Sargassum thunbergii (Mertens et Roth) Kuntze were carried out under controlled laboratory conditions. Growth characteristics of these germlings
grown under different temperatures (from 10 to 25°C), irradiances (from 9 to 88 μmol photons m−2 s−1), and under blue and white light conditions are described. The development of embryonic germlings follows the classic “8
nuclei 1 egg” type described for Sargassaceae. Fertilized eggs spent 5–6 h developing into multicellular germlings with abundant
rhizoids after fertilization. Under conditions of 20°C, 44 μmol photons m−2 s−1 and photoperiod of 12 h, young germlings with one or two leaflets reached 2–3 mm in length after 8 weeks. Temperature variations
(10, 15, 20, 25°C) under 88 μmol photons m−2 s−1 significantly influenced the growth rate within the first week, although this effect became less obvious after 8 weeks, especially
at 15 and 20°C. Variation in germling growth was highly significant under different irradiances (9, 18, 44, 88 μmol photons
m−2 s−1) at 25°C. Low temperature (10°C) reduced germling growth. Growth of germlings cultured under blue light was lower than in
white light. Optimal growth of these germlings occurred at 25°C and 44 μmol photons m−2 s−1. 相似文献
16.
Mechanism of peptide hydrolysis by co-catalytic metal centers containing leucine aminopeptidase enzyme: a DFT approach 总被引:1,自引:0,他引:1
Zhu X Barman A Ozbil M Zhang T Li S Prabhakar R 《Journal of biological inorganic chemistry》2012,17(2):209-222
In this density functional theory study, reaction mechanisms of a co-catalytic binuclear metal center (Zn1–Zn2) containing
enzyme leucine aminopeptidase for two different metal bridging nucleophiles (H2O and –OH) have been investigated. In addition, the effects of the substrate (l-leucine-p-nitroanilide → l-leucyl-p-anisidine) and metal (Zn1 → Mg and Zn2 → Co, i.e., Mg1–Zn2 and Mg1–Co2 variants) substitutions on the energetics of the mechanism
have been investigated. The general acid/base mechanism utilizing a bicarbonate ion followed by this enzyme is divided into
two steps: (1) the formation of the gem-diolate intermediate, and (2) the cleavage of the peptide bond. With the computed barrier of 17.8 kcal/mol, the mechanism
utilizing a hydroxyl nucleophile was found to be in excellent agreement with the experimentally measured barrier of 18.7 kcal/mol.
The rate-limiting step for reaction with l-leucine-p-nitroanilide is the cleavage of the peptide bond with a barrier of 17.8 kcal/mol. However, for l-leucyl-p-anisidine all steps of the mechanism were found to occur with similar barriers (18.0–19.0 kcal/mol). For the metallovariants,
cleavage of the peptide bond occurs in the rate-limiting step with barriers of 17.8, 18.0, and 24.2 kcal/mol for the Zn1–Zn2,
Mg1–Zn2, and Mg1–Co2 enzymes, respectively. The nature of the metal ion was found to affect only the creation of the gem-diolate intermediate, and after that all three enzymes follow essentially the same energetics. The results reported in this
study have elucidated specific roles of both metal centers, the nucleophile, indirect ligands, and substrates in the catalytic
functioning of this important class of binuclear metallopeptidases. 相似文献
17.
Soria F Ellenrieder G Oliveira GB Cabrera M Carvalho LB 《Applied microbiology and biotechnology》2012,93(3):1127-1134
α-l-Rhamnosidase from Aspergillus terreus was covalently immobilized on the following ferromagnetic supports: polyethylene terephthalate (Dacron-hydrazide), polysiloxane/polyvinyl
alcohol (POS/PVA), and chitosan. The powdered supports were magnetized by thermal coprecipitation method using ferric and
ferrous chlorides, and the immobilization was carried out via glutaraldehyde. The activity of the Dacron-hydrazide (0.53 nkat/μg
of protein) and POS/PVA (0.59 nkat/μg of protein) immobilized enzyme was significantly higher than that found for the chitosan
derivative (0.06 nkat/μg of protein). The activity–pH and activity–temperature profiles for all immobilized enzymes did not
show difference compared to the free enzyme, except the chitosan derivative that presented higher maximum temperature at 65 °C.
The Dacron-hydrazide derivative thermal stability showed a similar behavior of the free enzyme in the temperature range of
40–70 °C. The POS/PVA and chitosan derivatives were stable up to 60 °C, but were completely inactivated at 70 °C. The activity
of the preparations did not appreciably decrease after ten successive reuses. Apparent K
m of α-l-rhamnosidase immobilized on magnetized Dacron-hydrazide (1.05 ± 0.22 mM), POS/PVA (0.57 ± 0.09 mM), and chitosan (1.78 ± 0.24 mM)
were higher than that estimated for the soluble enzyme (0.30 ± 0.03 mM). The Dacron-hydrazide enzyme derivative showed better
performance than the free enzyme to hydrolyze 0.3% narigin (91% and 73% after 1 h, respectively) and synthesize rhamnosides
(0.116 and 0.014 mg narirutin after 1 h, respectively). 相似文献
18.
J. E. Gomes Júnior D. S. L. Souza R. M. Nascimento A. L. M. Lima J. A. T. Melo T. L. Rocha R. N. G. Miller O. L. Franco M. F. Grossi-de-Sa L. R. D. Abreu 《The protein journal》2010,29(3):188-194
A β-N-Acetylhexosaminidase (EC 3.2.1.52) was purified from hepatic extracts of Sotalia fluviatilis, order Cetacea. The protein was purified by using ammonium sulfate fractionation and four subsequent chromatographies (Biogel
A 1.5 m, Chitin, Deae-Biogel and hydroxyapatite resins). After these purification steps, the enzyme was purified 380.5-fold
with an 8.4% yield. The molecular mass (10 kDa) was estimated by SDS–PAGE and MALDI-TOF analysis. A Km of 2.72 mM and Vmax
9.5 × 10−6 μmol/(min.mg) were found for this enzyme, determined by p-nitrophenyl-β-d-hexosaminide substrate digestion. Optimal pH and temperature for β-N-Acetylhexosaminidase activity were 5.0 and 60 °C, respectively.
Enzyme activity was inhibited by sodium selenate (Na2SeO4), mercuric chloride (HgCl2) and sodium dodecyl sulfate (C12H25SO4Na), and activated by zinc, calcium, barium and lithium ions. Characterization of the β-N-Acetylhexosaminidase in Sotalia fluviatilis can be a basis for physiological studies in this species. 相似文献
19.
D.B. Light T.M. Mertins J.A. Belongia C.A. Witt 《The Journal of membrane biology》1997,158(3):229-239
We examined whether metabolites of arachidonic acid (AA) regulate K+ efflux during regulatory volume decrease (RVD) by mudpuppy red blood cells (RBCs). Volume regulation was inhibited by the
phospholipase A2 antagonists mepacrine (10 μm) and ONO-RS-082 (10 μm); the inhibitory effect of ONO-RS-082 was reversed by gramicidin (5 μm). Eicosatetraynoic acid (ETYA, 100 μm), a general antagonist of AA metabolism, also blocked RVD. In addition, volume regulation was inhibited by the lipoxygenase
pathway antagonist nordihydroguaiaretic acid (NDGA, 10 μm), the 5 lipoxygenase antagonists AA-861 (5 μm) and curcumin (20 μm), and by the 5-lipoxygenase activating protein inhibitor L-655,298 (5 μm). Inhibition by all four of these agents was reversed with gramicidin. In contrast, the 12- and 15-lipoxygenase pathway inhibitor
ethyl-3,4-dihydroxy-benzylidene-cyanoacetate (EDBCA, 1 μm) and the cytochrome P-450 monooxygenase pathway blocker ketoconazole (20 μm) had no effect. On the other hand, the cyclooxygenase pathway inhibitor aspirin (100 μm) slightly enhanced RVD. Consistent with these findings, a K+-selective whole cell conductance responsible for K+ efflux during cell swelling was inhibited by ONO-RS-082 (10 μm), NDGA (10 μm), AA-861 (5 μm), curcumin (20 μm), and l-655,298 (5 μm). In contrast, EDBCA (1 μm), ketoconazole (20 μm), and indomethacin (10 μm) did not block this whole cell conductance. These results indicate that a channel mediating K+ loss during RVD is regulated by a 5-lipoxygenase metabolite of arachidonic acid.
Received: 12 December 1996/Revised: 28 February 1997 相似文献
20.
Effect of Gibberellin on Growth, Protein Secretion, and Starch Accumulation in Maize Endosperm Suspension Cells 总被引:3,自引:0,他引:3
The physiologic effect of gibberellins (GA) in seed development is poorly understood. We examined the effect of gibberellic
acid (GA3) on growth, protein secretion, and starch accumulation in cultured maize (Zea mays L.) endosperm suspension cells. GA3 (5 and 30 μm) increased the fresh weight, dry weight, and protein content of the cultured cells, but the effect of GA3 at 50 μm was not significantly different. However, the protein content in the culture medium was increased by these three concentrations
of GA3. The effect of GA3 on the amount of cellular structural polysaccharides was not significant, but GA3 had a dramatic effect on the starch content. At 5 μm, GA3 caused an increase in the starch content, but at 50 μm the starch accumulation was reduced. Chlorocholine chloride (CCC), an inhibitor of GA biosynthesis, significantly increased
the starch content and decreased the structural polysaccharide content of the cultured cells. The effects of CCC at 500 μm on the starch and polysaccharide content were partially reversed by 5 μm GA3 applied exogenously. Based on these results we suggest that GA does not favor starch accumulation in the cell cultures and
that the addition of lower concentrations of GA3 in the medium may provide an improved balance among the endogenous GA in the cultured cells.
Received October 31, 1995; accepted March 25, 1997 相似文献