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81.
Purification of galectin-3 from ovine placenta: developmentally regulated expression and immunological relevance 总被引:1,自引:1,他引:1
Iglesias MM; Rabinovich GA; Ambrosio AL; Castagna LF; Sotomayor CE; Wolfenstein-Todel C 《Glycobiology》1998,8(1):59-65
Galectins, beta-galactoside-binding lectins, are extensively distributed in
the animal kingdom and share some basic molecular properties. Galectin-3, a
member of this family, is generally associated with differentiation,
morphogenesis, and metastasis. In this study, galectin-3 was isolated from
ovine placental cotyledons round the middle of the gestation period by
lactose extraction followed by affinity chromatography on lactosyl-agarose,
and separated from galectin-1 by size exclusion chromatography on a
Superose 12 column. Under native conditions this lectin behaved as a
monomer with an apparent molecular weight of approximately 29,000 and an
isoelectric point of 9.0. The partial amino acid sequence of the peptides
obtained by tryptic digestion of this protein followed by HPLC separation
showed striking homology with other members of the galectin-3 subfamily.
Furthermore, ovine placental galectin-3 exhibited specific mitogenic
activity toward rat spleen mononuclear cells. Besides, this protein
strongly reacted with a rabbit antiserum raised against a chicken galectin.
Results obtained by Western blot analysis showed that its expression was
greatly decreased in term placenta with respect to the middle of the
gestation period, suggesting a regulated expression throughout development.
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Nellis DF Giardina SL Janini GM Shenoy SR Marks JD Tsai R Drummond DC Hong K Park JW Ouellette TF Perkins SC Kirpotin DB 《Biotechnology progress》2005,21(1):221-232
Analytical methods optimized for micellar F5cys-MP-PEG(2000)-DPSE protein-lipopolymer conjugate are presented. The apparent micelle molecular weight, determined by size exclusion chromatography, ranged from 330 to 960 kDa. The F5cys antibody and conjugate melting points, determined by differential scanning calorimetry, were near 82 degrees C. Traditional methods for characterizing monodisperse protein species were inapplicable to conjugate analysis. The isoelectric point of F5cys (9.2) and the conjugate (8.9) were determined by capillary isoelectric focusing (cIEF) after addition of the zwitterionic detergent CHAPS to the buffer. Conjugate incubation with phospholipase B selectively removed DSPE lipid groups and dispersed the conjugate prior to separation by chromatographic methods. Alternatively, adding 2-propanol (29.4 vol %) and n-butanol (4.5 vol %) to buffers for salt-gradient cation exchange chromatography provided gentler, nonenzymatic dispersion, resulting in well-resolved peaks. This method was used to assess stability, identify contaminants, establish lot-to-lot comparability, and determine the average chromatographic purity (93%) for conjugate lots, described previously. The F5cys amino acid content was confirmed after conjugation. The expected conjugate avidity for immobilized HER-2/neu was measured by bimolecular interaction analysis (BIAcore). Mock therapeutic assemblies were made by conjugate insertion into preformed doxorubicin-encapsulating liposomes for antibody-directed uptake of doxorubicin by HER2-overexpressing cancer cells in vitro. Together these developed assays established that the manufacturing method as described in the first part of this study consistently produced F5cys-MP-PEG(2000)-DSPE having sufficient purity, stability, and functionality for use in preclinical toxicology investigations. 相似文献
84.
Howe GT; Bucciaglia PA; Hackett WP; Furnier GR; Cordonnier-Pratt MM; Gardner G 《Molecular biology and evolution》1998,15(2):160-175
The phytochrome photoreceptors play important roles in the photoperiodic
control of vegetative bud set, growth cessation, dormancy induction, and
cold-hardiness in trees. Interestingly, ecotypic differences in
photoperiodic responses are observed in many temperate- zone tree species.
Northern and southern ecotypes of black cottonwood (Populus trichocarpa
Torr. & Gray), for example, exhibit marked differences in the timing of
short-day-induced bud set and growth cessation, and these responses are
controlled by phytochrome. Therefore, as a first step toward determining
the molecular genetic basis of photoperiodic ecotypes in trees, we
characterized the phytochrome gene (PHY) family in black cottonwood. We
recovered fragments of one PHYA and two PHYB using PCR-based cloning and by
screening a genomic library. Results from Southern analyses confirmed that
black cottonwood has one PHYA locus and two PHYB loci, which we arbitrarily
designated PHYB1 and PHYB2. Phylogenetic analyses which included PHY from
black cottonwood, Arabidopsis thaliana and tomato (Solanum lycopersicum)
suggest that the PHYB/D duplications in these species occurred
independently. When Southern blots were probed with PHYC, PHYE, and PHYE
heterologous probes, the strongest bands that we detected were those of
black cottonwood PHYA and/or PHYB. These results suggest that black
cottonwood lacks members of the PHYC/F and PHYE subfamilies. Although black
cottonwood could contain additional PHY that are distantly related to known
angiosperm PHY, our results imply that the PHY family of black cottonwood
is less complex than that of other well-characterized dicot species such as
Arabidopsis and tomato. Based on Southern analyses of five black cottonwood
genotypes representing three photoperiodic ecotypes, substantial
polymorphism was detected for at least one of the PHYB loci but not for the
PHYA locus. The novel character of the PHY family in black cottonwood, as
well as the differences in polymorphism we observed between the PHYA and
PHYB subfamilies, indicates that a number of fundamental macro- and
microevolutionary questions remain to be answered about the PHY family in
dicots.
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Exosomes secreted from mutant‐HIF‐1α‐modified bone‐marrow‐derived mesenchymal stem cells attenuate early steroid‐induced avascular necrosis of femoral head in rabbit 下载免费PDF全文
89.
Annemarie MM Vlaar Tjerk de Nijs Marinus JPG van Kroonenburgh Werner H Mess Ania Winogrodzka Selma C Tromp Wim EJ Weber 《BMC neurology》2008,8(1):42
Background
Transcranial duplex sonography (TCD) of the substantia nigra has emerged as a promising, non-invasive tool to diagnose idiopathic Parkinson's disease (IPD). However, its diagnostic accuracy in patients with undefined parkinsonism remains to be determined. 相似文献90.
Sterigmatocystin (STG) is a toxic metabolite produced by severalAspergillus species. Because of its toxic and carcinogenic properties the occurrence of STG in food is considered to represent a potential
hazard to man. The present study was designed to investigate following points:
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1 | A survey of STG incidence in Ras cheese on local markets. Ras cheese samples were collected from Cairo, Giza and Kalubia governorates. Thirty five percent of the samples contained the toxin with a mean value of 22.23 μg /kg |
2 | Fate of STG contaminating milk during Ras cheese processing. Milk was artificially contaminated with 125 μg/kg and processed into Ras cheese. Eighty percent of the toxin was distributed into the curd and 20% into the whey. Cheese ripening effected toxin content and the effect was temperature dependent. At 6°C: toxin concentration was slightly affected; at 20°C the toxin was reduced by 16% after 90 days when low toxin concentration was used. |
3 | Formation of STG byA versicolor mold on Ras cheese. Ras cheese blocks were contaminated with spores of the mold. Toxin production started after 45 days of ripening and reached a maximum at 90 days and then declined. Cow’s milk favoured toxin production over buffaloe’s. Aged cheese inhibited toxin production. |