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1.
A lectin with antifungal and mitogenic activities from red cluster pepper (<Emphasis Type="Italic">Capsicum frutescens</Emphasis>) seeds 总被引:1,自引:0,他引:1
A monomeric mannose/glucose-binding lectin, with a molecular mass of 29.5 kDa and an N-terminal sequence GQRELKL showing resemblance
to that of the lectin-like oxidized low-density lipoprotein receptor from the rabbit, has been isolated from the seeds of
red cluster pepper Capsium frutescens L. var. fasciculatum. The protocol involved anion exchange chromatography on diethylamino ethanol-cellulose and Q-Sepharose and fast protein liquid
chromatography on Mono Q. Its hemagglutinating activity toward rabbit erythrocytes was inhibited by d-mannose and glucose, specifically. The activity was stable from 0 to 40°C, reached a maximum at pH 7 and 8, and was potentiated
by Ca2+ and Mn2+ ions. The lectin showed strong mitogenic activity toward spleen cells isolated from BALB/c mice. The mitogenic activity,
which reached a peak at a lectin concentration of 0.27 μM, was inhibited specifically by d(+)-mannose. The lectin was capable of inhibiting the germination of Aspergillus flavus and Fusarium moniliforme spores and hyphal growth in the two fungi. 相似文献
2.
An N-acetyl-d-lactosamine (LacNAc) specific lectin from tubers of Alocasia cucullata was purified by affinity chromatography on asialofetuin-linked amino activated silica. The pure lectin showed a single band
in SDS-PAGE at pH 8.8 and was a homotetramer with a subunit molecular mass of 13.5 kDa and native molecular mass of 53 kDa.
It was heat stable up to 55 °C for 15 min and showed optimum hemagglutination activity from pH 2 to 11. The lectin was affected
by denaturing agents such as urea (2 m), thiourea (2 m) and guanidine–HCl (0.5 m) and did not require Ca2+ and Mn2+ for its activity. It was a potent mitogen at 10 μg/ml towards human peripheral blood mononuclear cells with 50% growth inhibitory
potential towards SiHa (human cervix ) cancer cell line at 100 μg/ml. 相似文献
3.
Dresch RR Lerner CB Mothes B Trindade VM Henriques AT Vozári-Hampe MM 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2012,161(4):365-370
Lectin II from the marine sponge Axinella corrugata (ACL-II) was purified by affinity chromatography on rabbit erythrocytic stroma incorporated into a polyacrylamide gel, followed by gel filtration on Ultrogel AcA 44 column. Purified ACL-II is a lectin with an Mr of 80 kDa and 78 kDa, estimated by SDS-PAGE and by FPLC on Superose 12 HR column, respectively. ACL-II mainly agglutinates native rabbit erythrocytes and this hemagglutinating activity is independent of Ca2 +, Mg2 + and Mn2 +, but is inhibited by d-galactose, chitin and N-acetyl derivatives, with the exception of GalNAc. ACL-II is stable for up to 65 °C for 30 min, with a better stability at a pH range of 2 to 6. In contrast, ACL-I displays a strong mitogenic and cytotoxic effect. 相似文献
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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.
Michelin M Ruller R Ward RJ Moraes LA Jorge JA Terenzi HF Polizeli Mde L 《Journal of industrial microbiology & biotechnology》2008,35(1):17-25
An extracellular glucoamylase produced by Paecilomyces variotii was purified using DEAE-cellulose ion exchange chromatography and Sephadex G-100 gel filtration. The purified protein migrated
as a single band in 7% PAGE and 8% SDS-PAGE. The estimated molecular mass was 86.5 kDa (SDS-PAGE). Optima of temperature and
pH were 55 °C and 5.0, respectively. In the absence of substrate the purified glucoamylase was stable for 1 h at 50 and 55 °C,
with a t
50 of 45 min at 60 °C. The substrate contributed to protect the enzyme against thermal denaturation. The enzyme was mainly activated
by manganese metal ions. The glucoamylase produced by P. variotii preferentially hydrolyzed amylopectin, glycogen and starch, and to a lesser extent malto-oligossacarides and amylose. Sucrose,
p-nitrophenyl α-d-maltoside, methyl-α-d-glucopyranoside, pullulan, α- and β-cyclodextrin, and trehalose were not hydrolyzed. After 24 h, the products of starch hydrolysis,
analyzed by thin layer chromatography, showed only glucose. The circular dichroism spectrum showed a protein rich in α-helix.
The sequence of amino acids of the purified enzyme VVTDSFR appears similar to glucoamylases purified from Talaromyces emersonii and with the precursor of the glucoamylase from Aspergillus oryzae. These results suggested the character of the enzyme studied as a glucoamylase (1,4-α-d-glucan glucohydrolase). 相似文献
9.
Ponpimol Tipthara Polkit Sangvanich Marcus Macth Amorn Petsom 《Journal of Plant Biology》2007,50(2):167-173
A mannose-binding lectin was isolated from rhizomes of the medicinal plantCurcuma zedoaria. We used extraction with 20 mM phosphate buffer, ammonium sulfate precipitation, ion exchange chromatography on Q-Sepharose,
gel filtration chromatography on Superdex 75, and reverse-phase HPLC. The purified lectin yielded a single band on SDS-PAGE
that corresponded to a molecular mass of 13 kDa. This lectin exhibited hemagglutinating activity toward rabbit erythrocytes,
which could be inhibited by mannose only. The lectin was digested with trypsin and its digests were analyzed using MALDI-TOF/TOF.
Partial amino acid sequences were obtained from tandem mass spectra via automatedde novo sequencing, and were then identified by MS-BLAST homology searches to enable recognition of related proteins in other species.
Inferred peptide sequences exhibited similarity to a mannose-binding lectin fromEpipactis helleborine, a member of the Orchidaceae. 相似文献
10.
Dresch RR Zanetti GD Lerner CB Mothes B Trindade VM Henriques AT Vozári-Hampe MM 《Comparative biochemistry and physiology. Toxicology & pharmacology : CBP》2008,148(1):23-30
The lectin from the marine sponge Axinella corrugata (ACL-I) was purified by affinity chromatography on rabbit erythrocytic stroma incorporated into a polyacrylamide gel followed by gel filtration on Ultrogel AcA 44 column. Purified ACL-I is a hexameric glycoprotein with a Mr of 82.3 kDa estimated by SDS-PAGE and 78.5 kDa by FPLC on Superose 12 HR column. The pI of lectin is 6.3 and ACL-I is constituted of 13.9 kDa similar subunits some of them linked by disulphide bridges. This lectin agglutinates native rabbit, goat and dog erythrocytes and in less extent human erythrocytes. The hemagglutinating activity is independent of Ca(2+), Mg(2+) and Mn(2+), but it is strongly inhibited by carbohydrates containing N-acetyl groups. ACL-I is stable up to 70 degrees C for 30 min, with optimum pH between 7 and 8, and it is also resistant to enzymatic proteolysis in vitro. In the presence of reducing or denaturant agents, the lectin activity decreases. ACL-I displays chemotactic effect on rat neutrophil in vitro which is inhibited by N-acetyl-d-glucosamine. 相似文献
11.
Linda S. M. Ooi Wing-Shan Ho Karry L. K. Ngai Li Tian Paul K. S. Chan Samuel S. M. Sun Vincent E. C. Ooi 《Journal of biosciences》2010,35(1):95-103
A mannose-binding lectin (Narcissus tazetta lectin [NTL]) with potent antiviral activity was isolated and purified from the bulbs of the Chinese daffodil Narcissus tazetta var. chinensis, using ion exchange chromatography on diethylaminoethyl (DEAE)-cellulose, affinity chromatography on mannose-agarose and
fast protein liquid chromatography (FPLC)-gel filtration on Superose 12. The purified lectin was shown to have an apparent
molecular mass of 26 kDa by gel filtration and 13 kDa by SDS-PAGE, indicating that it is probably a dimer with two identical
subunits. The cDNA-derived amino acid sequence of NTL as determined by molecular cloning also reveals that NTL protein contains
a mature polypeptide consisting of 105 amino acids and a C-terminal peptide extension. Three-dimensional modelling study demonstrated
that the NTL primary polypeptide contains three subdomains, each with a conserved mannose-binding site. It shows a high homology
of about 60%–80% similarity with the existing monocot mannose-binding lectins. NTL could significantly inhibit plaque formation
by the human respiratory syncytial virus (RSV) with an IC50 of 2.30 μg/ml and exhibit strong antiviral properties against influenza A (H1N1, H3N2, H5N1) and influenza B viruses with
IC50 values ranging from 0.20 μg/ml to 1.33 μg/ml in a dose-dependent manner. It is worth noting that the modes of antiviral action
of NTL against RSV and influenza A virus are significantly different. NTL is effective in the inhibition of RSV during the
whole viral infection cycle, but the antiviral activity of NTL is mainly expressed at the early stage of the viral cycle of
influenza A (H1N1) virus. NTL with a high selective index (SI=CC50/IC50≥141) resulting from its potent antiviral activity and low cytotoxicity demonstrates a potential for biotechnological development
as an antiviral agent. 相似文献
12.
A mannose-binding lectin was isolated from leaves of the Chinese daffodil Narcissus tazetta (family Amaryllidaceae) using a procedure that comprised extraction with aqueous buffer, ammonium sulfate precipitation, ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel Blue gel and mannose-agarose, and FPLC-gel filtration on Superose 12. The lectin was adsorbed on mannose-agarose and unadsorbed on DEAE-cellulose and Affi-gel Blue gel. It was an unglycosylated homodimer with a molecular mass of 26 kDa. Analysis of the N-terminal sequence of the N. tazetta lectin revealed considerable homology to lectins from the daffodil Narcissus pseudonarcissus, the snowdrop Galanthus nivalis (family Amaryllidaceae), the tulip Tulipa, and Kidachi aloe Aloe arborescens (family Liliaceae), and the orchid lectins (family Orchidaceae). The most striking likeness exists among the Amaryllidaceae lectins. The N. tazetta lectin exhibits hemagglutinating activity toward rabbit erythrocytes. 相似文献
13.
d-Xylose/d-glucose isomerases from two strains, a newly isolated strain, Paenibacillus sp., and from Alcaligenes ruhlandii are described herein. The enzymes were purified to apparent homogeneity. Both of these d-xylose isomerases are homotetramers with relative subunit molecular masses of 45 000 and 53 000, respectively, as estimated
by sodium dodecylsulphate-polyacrylamide gel electrophoresis. The native molecular masses determined on Superose 12 gel chromatography
are 181 kDa for the enzyme from Paenibacillus sp. and 199 kDa for that from A. ruhlandii. The activity of both enzymes shows a requirement for divalent metal ions; the d-xylose isomerase from Paenibacillus sp. has the highest activity with Mn2+, while the enzyme from A. ruhlandii prefers Mg2+. Both enzymes also accept Co2+ with a somewhat lower efficiency, while Cu2+ inhibits the enzyme reaction. The binding of the metal ions obeys a biphasic characteristic, indicating the presence of two
non-identical binding sites per subunit. d-Glucose is converted to d-fructose at a rate that is two- to three-fold slower than for the d-xylose isomerisation. d-Xylitol and d-lyxose are competitive inhibitors of both enzymes. Both enzymes have a pH optimum between 6.5 and 7.0, and they are active
up to 60 °C. The enzyme from Paenibacillus sp. retained 50% of its activity after 4 days at 55 °C, whereas that from A. ruhlandii still retained 50% of its activity after 6 days at 55 °C. Polyacrylamide entrapment and immobilisation to both controlled
pore glass and cyanogen-bromide-activated Sepharose were achieved for both enzymes with high efficiency.
Received: 14 May 1998 / Received last revision: 29 July 1998 / Accepted: 29 July 1998 相似文献
14.
Masako Higuchi Yutaka Ohtani Kazuo Iwai 《Bioscience, biotechnology, and biochemistry》2013,77(7):1847-1853
Winged bean acidic lectin was purified by DEAE-Sephadex A-50 and affinity chromatography on N-acetylgalactosamine-agarose gel. The purified lectin was a glycoprotein homogeneous on polyacrylamide gel electrophoresis, isoelectric focusing, and gel filtration. The molecular weight of the lectin was 52,000 by gel filtration, and SDS-polyacrylamide gel electrophoresis gave a single component of molecular weight of 27,000. Its isoelectric point was 5.5. The acidic lectin was rich in acidic amino acids, and contained 2mol of methionine but no cystine. It also agglutinated both trypsinized and untreated human erythrocytes (types A, B, AB and O), but not rabbit erythrocytes. The hemagglutination was inhibited by d-galactose and related sugars. Modification of the acidic lectin with N-bromosuccinimide caused a concomitant loss of the hemagglutinating activity with oxidation of tryptophan residue. The acidic lectin was immunologically different from the purified winged bean basic lectin by double immunodiffusion using antiserum raised against the basic lectin. 相似文献
15.
Min Gui Jung Key Pyoung Lee Han-Gu Choi Sung-Ho Kang Tatyana A. Klochkova Jong Won Han Gwang Hoon Kim 《Journal of applied phycology》2010,22(6):793-802
Bryohealin is a lectin involved in the wound-healing process of the marine green alga Bryopsis plumosa. In the previous purification study, it has been shown that lectin was composed of two identical subunits of 27 kDa, cross-linked
by disulfide bond, and showed binding specificity to N-acetyl-d-glucosamine and N-acetyl-d-galactosamine (GlcNAc and GalNAc, respectively). To determine if the lectin recognize the two different sugars at the same
binding domain, the carbohydrate binding sites of Bryohealin was analyzed using chromatography and chemical modification methods.
Results showed that the same binding site of the lectin was responsible for the recognition of two sugars, GalNAc as well
as GlcNAc. Chemical modification studies showed that hemagglutinating activities of Bryohealin were not affected by modification
of histidine, tryptophan, aspartic acid, and glutamic acid. When arginine residues were modified with 1,2-cyclohexanedione,
the activity of Bryohealin rapidly decreased. The sugar binding sites remained intact when the lectin was treated with inhibitory
sugars (0.2 M GalNAc and/or GlcNAc) prior to 1,2-cyclohexanedione treatment. The sugar binding domain of Bryohealin was predicted
from the MALDI-TOF analysis and the full cDNA sequence of the lectin gene. 相似文献
16.
An α-D-galactose-specific lectin from the seeds of jack fruit (Artocarpus integra) has been isolated in pure form by affinity chromatography on immobilised guar gum (a galactomannan). The lectin is shown
to be a glycoprotein containing 3% carbohydrate and having a molecular weight of 39,500 as determined by gel filtration. Sodium
dodecyl sulphate gel electrophoresis revealed a single polypeptide of 10,500 dalton, indicating that the native lectin is
a tetrarner of identical subunits. The hemagglutinating activity of the lectin towards erythrocytes of all blood groups is
found to be the same. 相似文献
17.
A lectin present in seeds of Clitoria ternatea agglutinated trypsin-treated human B erythrocytes. The sugar specificity assay indicated that lectin belongs to Gal/Gal NAc-specific
group. Hence the lectin, designated C. ternatea agglutinin (CTA), was purified by the combination of acetic acid precipitation, salt fractionation and affinity chromatography.
HPLC gel filtration, SDS-polyacrylamide gel electrophoresis and mass spectrometry indicated that the native lectin is composed
of two identical subunits of molecular weight 34.7 kDa associated by non covalent bonds. The N-terminal sequence of CTA shared
homology with Glycine max and Pisum sativum. Complete sequence was also found to be homologous to S-64 protein of Glycine max, suggesting that CTA probably exhibits both hemagglutination and probably sugar uptake activity. The carbohydrate binding
specificity of the lectin was investigated by quantitative turbidity measurements, and percent inhibition assays. Based on
these assays, we conclude that CTA binds β-d-galactosides, and also may has an extended specificity towards non-reducing terminal Neu5Acα2,6Gal. 相似文献
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19.
Ng TB Yu YL Chu KT 《Comparative biochemistry and physiology. Toxicology & pharmacology : CBP》2002,133(3):453-460
A novel mannose- and glucose-specific lectin with high hemagglutinating activity was isolated from seeds of the Chinese chestnut Castanea mollisima. The lectin possessed a molecular mass of 140 kDa and was made up of two subunits, one with a molecular mass of 31 kDa and another with a molecular mass of 32 kDa. They exhibited substantial homology in N-terminal sequence to the storage protein legumin. The lectin was unstable in the presence of acid and alkali and at temperatures above 50 degrees C, but it was unaffected by various salts. The lectin was purified with a procedure involving ion exchange chromatography on CM-Sepharose, Q-Sepharose and Resource Q and gel filtration on Superose 12. 相似文献
20.
Sialidase secreted by the urease-positiveClostridium sordellii strain G12 was isolated from culture medium and purified to apparent homogeneity as estimated by Fast Protein Liquid Chromatography (FPLC) and sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE). For this purpose, ion-exchange chromatography, gel filtration, isoelectric focusing, and FPLC on ion-exchange resin and gel filtration materials were used. The sialidase was purified 159 300-fold from 5 l of culture medium, yielding 9 g of enzyme protein with a specific activity of 480 U/mg. For the denatured (SDS-PAGE) and native (FPLC) sialidase relative molecular masses of 40 000 and 38 500 Da, respectively, were estimated. The substrate specificity, kinetic data, and pH-optimum of the enzyme are similar to those of other bacterial sialidases. The influences of salt or serum proteins on enzyme activity are of interest.Abbreviations MU-Neu5Ac
4-methylumbelliferyl -d-N-acetylneuraminic acid
- Ganglioside GD1a
IV3NeuAc, ll3NeuAc-GgOse4Cer
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid 相似文献