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61.
Epitopes, structural domains, and asymmetry of amino acid residues in SS-B/La nuclear protein 总被引:21,自引:0,他引:21
E K Chan A M Francoeur E M Tan 《Journal of immunology (Baltimore, Md. : 1950)》1986,136(10):3744-3749
SS-B/La is a conserved cellular phosphoprotein of 46 to 48 KD that is the target antigen of autoantibodies in sera of patients with Sjogren's syndrome and systemic lupus erythematosus. SS-B/La is also known to be associated with certain small cellular and viral RNA, including adenovirus VAI and VAII RNA. Two relatively protease-resistant domains (X and Y) were defined in SS-B from HeLa cells by using human autoantibodies as reagents. Domain X, a methionine-containing nonphosphorylated 28 KD polypeptide, was found to be resistant to partial digestion with six different proteases. Similar domains were also found in calf and rabbit SS-B. Domain Y, a 23 KD polypeptide, was detected after limited digestion with S. aureus V8 and trypsin. This domain contained little if any methionine, but all the detectable phosphorylated amino acids. Among 16 anti-SS-B sera tested by immunoblotting, 11 (69%) were reactive with both domains, three (19%) only with domain X, and two (13%) only with domain Y. These results showed that there are at least two distinct antigenic epitopes on the 46 to 48 KD SS-B/La protein, each located on a separate structural domain. The asymmetric distribution of methionine and phosphorylated amino acid residues in SS-B/La show striking similarity to the two reported domains of the adenovirus 72 KD DNA-binding protein, and raises questions concerning functional similarities that await investigation. 相似文献
62.
An auxiliary protein for DNA polymerase-delta from fetal calf thymus 总被引:62,自引:0,他引:62
An auxiliary protein which affects the ability of calf thymus DNA polymerase-delta to utilize template/primers containing long stretches of single-stranded template has been purified to homogeneity from the same tissue. The auxiliary protein coelutes with DNA polymerase-delta on DEAE-cellulose and phenyl-agarose chromatography but is separated from the polymerase on phosphocellulose chromatography. The physical and functional properties of the auxiliary protein strongly resemble those of the beta subunit of Escherichia coli DNA polymerase III holoenzyme. A molecular weight of 75,000 has been calculated from a sedimentation coefficient of 5.0 s and a Stokes radius of 36.5 A. A single band of 37,000 daltons is seen on sodium dodecyl sulfate gel electrophoresis, suggesting that the protein exists as a dimer of identical subunits. The purified protein has no detectable DNA polymerase, primase, ATPase, or nuclease activity. The ability of DNA polymerase-delta to replicate gapped duplex DNA is relatively unaffected by the presence of the auxiliary protein, however, it is required to replicate templates with low primer/template ratios, e.g. poly(dA)/oligo(dT) (20:1), primed M13 DNA, and denatured calf thymus DNA. The auxiliary protein is specific for DNA polymerase-delta; it has no effect on the activity of calf thymus DNA polymerase-alpha or the Klenow fragment of E. coli DNA polymerase I with primed homopolymer templates. Although the auxiliary protein does not bind to either single-stranded or double-stranded DNA, it does increase the binding of DNA polymerase-delta to poly(dA)/oligo(dT), suggesting that the auxiliary protein interacts with the polymerase in the presence of template/primer, stabilizing the polymerase-template/primer complex. 相似文献
63.
The populations of native male adult oriental fruit fly Dacus dorsalis (Hendel ) and artocarpus fruit fly D. umbrosus (F.) in two selected site (BU and SD) were estimated weekly by the capture-recapture technique using live traps baited with methyl eugenol. In BU where many varieties of fruit trees were grown, the estimated population densities of D. dorsalis were between 980 and 3100 male flies per ha between May and July, 1984. During the same period, in SD where there were fewer number and varieties of fruit trees, the estimated population densities were between 300 and 1000 flies per ha. The estimated population densities of D. umbrosus over the same period were between 570 and 1290 flies per ha in BU; and between 5 and 95 flies per ha in SD. Of a total 6828 marked D. dorsalis flies released only one fly (released 6 weeks earlier in BU) was caught in a different site. 相似文献
64.
Cloning and expression of a Streptococcus cremoris proteinase in Bacillus subtilis and Streptococcus lactis. 总被引:10,自引:9,他引:1 下载免费PDF全文
Previously, curing experiments suggested that plasmid pWV05 (17.5 megadaltons [Md]) of Streptococcus cremoris Wg2 specifies proteolytic activity. A restriction enzyme map of pWV05 was constructed, the entire plasmid was subcloned in Escherichia coli with plasmids pBR329 and pACYC184. A 4.3-Md HindIII fragment could not be cloned in an uninterrupted way in E. coli but could be cloned in two parts. Both fragments showed homology with the 9-Md proteinase plasmid of S. cremoris HP. The 4.3-Md HindIII fragment was successfully cloned in Bacillus subtilis on plasmid pGKV2 (3.1 Md). Crossed immunoelectrophoresis of extracts of B. subtilis carrying the recombinant plasmid (pGKV500; 7.4 Md) showed that the fragment specifies two proteins of the proteolytic system of S. cremoris Wg2. PGKV500 was introduced in a proteinase-deficient Streptococcus lactis strain via protoplast transformation. Both proteins were also present in cell-free extracts of S. lactis(pGKV500). In S. lactis, pGKV500 enables the cells to grow normally in milk with rapid acid production, indicating that the 4.3-Md HindIII fragment of plasmid pWV05 specifies the proteolytic activity of S. cremoris Wg2. 相似文献
65.
66.
Exclusive leukotriene C4 synthesis by purified human eosinophils induced by opsonized zymosan 总被引:4,自引:0,他引:4
Purified human eosinophils were challenged with N-formyl-methionyl-leucyl-phenylalanine, leukotriene B4, platelet-activating-factor, valyl-glycyl-seryl-glutamic acid, phorbol myristate acetate, zymosan, opsonized zymosan and the calcium ionophore A23187 to induce leukotriene synthesis. Reversed-phase high performance liquid chromatography analysis demonstrated the almost exclusive synthesis of leukotriene C4 by eosinophils of 11 healthy donors after challenge with opsonized zymosan [(22 +/- 4) X 10(6) molecules LTC4/cell, mean +/- SE] or the calcium ionophore A23187 [(54 +/- 7) X 10(6) molecules LTC4/cell, mean +/- SE]. The other agents were not capable of inducing leukotriene formation. When in addition to opsonized zymosan N-formyl-methionyl-leucyl-phenylalanine or platelet-activating factor were added a significant increase of the leukotriene C4 synthesis by eosinophils was observed. These results suggest that eosinophils might be triggered to produce considerable amounts of the spasmogenic leukotriene C4 in vivo by C3b- and/or IgG-mediated mechanisms e.g. phagocytosis. 相似文献
67.
J. D. Tan 《Journal of fish biology》1985,26(6):657-668
The pituitary gland of the milkfish, Chanos chanos , was studied at different stages of sexual maturation and spawning. Consecutive median sagittal sections were treated with a range of stains to demonstrate the different cell types and regions. The milkfish pituitary consists of a neural component, the neurohypophysis, and an epithelial component, the adenohypophysis, which in turn consists of three regions: the rostral pars distalis (RPD), proximal pars distalis (PPD), and pars intermedia (PI). However, unlike most teleosts, the pituitary gland of the milkfish is encased in a bony chamber, has dorsal and ventral lobes and extends anteriorly from its point of origin at the base of the brain. PAS (+) basophils are found in all regions of the adenohypophysis, but mostly in the proximal pars distalis. These cells undergo hypertrophy and hyperplasia during sexual maturation, shrinkage and degranulation during spawning. 相似文献
68.
C W Chi D X Zhu N Q Lin L X Xu F L Tan L X Wang 《Biological chemistry Hoppe-Seyler》1985,366(9):879-885
After reduction and alkylation of the disulfide bonds of the proteinase inhibitor B from the root of the arrowhead (Sagittaria sagittifolia L.) followed by CNBr cleavage three peptide fragments with 68, 62 and 11 amino-acid residues could be separated on DEAE-Sepharose CL-6B. The peptides or the inhibitor itself were further specifically cleaved either by trypsin or by the mixture of (CH3)2SO/HCl/HBr at the arginyl- and the tryptophyl-peptide bond, respectively. The complete amino-acid sequences of the peptides were determined by manual solid phase DABITC/PITC double coupling micro-method and the primary structure of the arrowhead inhibitor B consisting of 141 amino-acid residues was then elucidated. Twenty pairs of amino-acid residues are repeated in the molecule of this inhibitor, three of these pairs even occur three times. The possible locations of the reactive sites are discussed. On the basis of sequence comparisons between this inhibitor and all other serine proteinase inhibitors the arrowhead inhibitor may belong to a new family. 相似文献
69.
A method is described for the preparation of both Factor X1 and Factor X2 from citrated bovine blood. The proteins from the plasma were first adsorbed on barium citrate by adding barium chloride solution. The precipitate formed was stirred with citrate/NaOH pH 6.9 buffer; barium and other clotting factors were removed by adding ammonium sulphate (up to 30% saturation) to the suspension. The Factor X was then precipitated by 65% ammonium sulphate, after resolution in citrate buffer chromatographed on DEAE-Sephadex and purified by rechromatography on DEAE-Sephadex and DEAE-Sepharose, respectively. This yielded Factor X1 and Factor X2 with respective purifications of about 16 000 and 24 000-fold that of the plasma. The apparent molecular mass of both Factor X1 and Factor X2 was 55 kDa as estimated by the sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Factor X2 had a higher specific biological activity of about 340 000 units/mg compared to that of Factor X1 of about 230 000 units/mg. 相似文献
70.
Cloning and expression of a Streptococcus cremoris proteinase in Bacillus subtilis and Streptococcus lactis 总被引:10,自引:0,他引:10
J Kok J M van Dijl J M van der Vossen G Venema 《Applied and environmental microbiology》1985,50(1):94-101
Previously, curing experiments suggested that plasmid pWV05 (17.5 megadaltons [Md]) of Streptococcus cremoris Wg2 specifies proteolytic activity. A restriction enzyme map of pWV05 was constructed, the entire plasmid was subcloned in Escherichia coli with plasmids pBR329 and pACYC184. A 4.3-Md HindIII fragment could not be cloned in an uninterrupted way in E. coli but could be cloned in two parts. Both fragments showed homology with the 9-Md proteinase plasmid of S. cremoris HP. The 4.3-Md HindIII fragment was successfully cloned in Bacillus subtilis on plasmid pGKV2 (3.1 Md). Crossed immunoelectrophoresis of extracts of B. subtilis carrying the recombinant plasmid (pGKV500; 7.4 Md) showed that the fragment specifies two proteins of the proteolytic system of S. cremoris Wg2. PGKV500 was introduced in a proteinase-deficient Streptococcus lactis strain via protoplast transformation. Both proteins were also present in cell-free extracts of S. lactis(pGKV500). In S. lactis, pGKV500 enables the cells to grow normally in milk with rapid acid production, indicating that the 4.3-Md HindIII fragment of plasmid pWV05 specifies the proteolytic activity of S. cremoris Wg2. 相似文献