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11.
Characterization and expression of two avirulence genes cloned from Pseudomonas syringae pv. glycinea. 总被引:12,自引:5,他引:7
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Two avirulence genes, avrB and avrC, from race 0 of Pseudomonas syringae pv. glycinea, were sequenced and found to encode single protein products of 36 and 39 kilodaltons, respectively. The proteins had neither recognizable signal peptide sequences nor significant stretches of hydrophobic amino acids that might indicate membrane association. Both avrB and avrC had relatively low position 3 and overall G+C contents, which suggests that they may have been recently introduced into P. syringae pv. glycinea. The deduced amino acid sequences of the proteins encoded by avrB and avrC shared 42% identical amino acids. However, when introduced into race 4 of P. syringae pv. glycinea, each gene directed a unique pattern of hypersensitive reactions on several differential soybean cultivars. The avrC protein was overproduced in Escherichia coli cells and deposited as insoluble inclusion bodies in the cell cytoplasm. The avrC protein could be solubilized with urea-octyl glucoside treatment, but neither the solubilized protein nor the intact inclusion bodies elicited a hypersensitive reaction in soybean leaves. 相似文献
12.
Y Iwatani N Amino H Tamaki M Aozasa O Kabutomori M Mori O Tanizawa K Miyai 《Endocrinologia japonica》1988,35(3):447-453
In order to elucidate the mechanism of postpartum aggravation of autoimmune thyroid disease (AITD), we serially examined the change in the proportion of peripheral large granular lymphocytes (LGL), which have activities of NK, K and/or cytotoxic T cells, in their postpartum period. Within 6 months postpartum, the percentage of LGL increased transiently in patients with AITD who remained euthyroid, or developed destructive thyrotoxicosis and/or hypothyroidism due to thyroiditis and even in normal controls. These changes in the LGL percentage were more obvious in the patients who had marked postpartum thyroid dysfunction. In contrast, we did not find a definite increase in the LGL percentage within 6 months postpartum in patients with Graves' disease who relapsed into Graves' thyrotoxicosis. These deta suggest that the increase in LGL in the postpartum period may be related to the induction of postpartum destructive thyrotoxicosis and/or hypothyroidism in AITD. 相似文献
13.
I (C3b/C4b inactivator) typing by agarose gel isoelectric focusing and immunoblotting technique 总被引:1,自引:0,他引:1
Genetic polymorphism of I (C3b/C4b inactivator) was studied by the method of agarose gel isoelectric focusing followed by an immunoblotting technique. Serum or plasma samples were pretreated with neuraminidase. The method is rapid, and gives the simple and reliable patterns of I. The allele frequencies calculated from healthy Japanese individuals living in the western part of Japan were: IF* A = 0.126 and IF*B = 0.874. 相似文献
14.
Peptidoglycan synthetic activities in membranes of Escherichia coli caused by overproduction of penicillin-binding protein 2 and rodA protein 总被引:27,自引:0,他引:27
F Ishino W Park S Tomioka S Tamaki I Takase K Kunugita H Matsuzawa S Asoh T Ohta B G Spratt 《The Journal of biological chemistry》1986,261(15):7024-7031
Penicillin-binding protein (PBP)-2 and the RodA protein are known to function in determining the rod shape of Escherichia coli cells. Peptidoglycan biosynthetic reactions that required these two proteins were demonstrated in the membrane fraction prepared from an E. coli strain that overproduced both of these two proteins and which lacked PBP-1B activity (the major peptidoglycan synthetase activity in the normal E. coli membranes). The cross-linked peptidoglycan was synthesized from UDP-N-acetylmuramylpentapeptide and UDP-N-acetylglucosamine in the presence of a high concentration of cefmetazole that inhibited all of PBPs except PBP-2. The peptidoglycan was synthesized via a lipid intermediate and showed up to 30% cross-linking. The cross-linking reaction was strongly inhibited by the amidinopenicillin, mecillinam, and by other beta-lactam antibiotics that have a high affinity for PBP-2, but not by beta-lactams that had very low affinity for PBP-2. The formation of peptidoglycan required the presence of high levels of both PBP-2 and the RodA protein in the membranes, but it is unclear which of the two proteins was primarily responsible for the extension of the glycan chains (transglycosylation). However, the sensitivity of the cross-linking reaction to specific beta-lactam antibiotics strongly suggested that it was catalyzed by PBP-2. The transglycosylase activity of the membranes was sensitive to enramycin and vancomycin and was unusual in being stimulated greatly by a high concentration of a chelating agent. 相似文献
15.
Purification and properties of mitochondrial malate dehydrogenase from unfertilized eggs of the sea urchin, Anthocidaris crassispina 总被引:1,自引:0,他引:1
Purification and characterization of mitochondrial malate dehydrogenase [EC 1.1.1.37] from unfertilized eggs of the sea urchin, Anthocidaris crassispina, are described. The purification method consisted of dextran sulfate fractionation, Blue Dextran Sepharose chromatography, Phenyl-Sepharose hydrophobic chromatography and DEAE-cellulose chromatography. The enzyme was purified 771-fold with a 7% yield from the crude extract. The purified enzyme appeared homogeneous on polyacrylamide gel electrophoresis under both native and denatured conditions. After incubation at 45 degrees C for 50 min, the enzyme lost about 90% of its activity. In the presence of NADH, however, the enzyme was protected against the heat denaturation. The native enzyme had a molecular weight of about 65,000 and probably consisted of two identical subunits. In the reduction of oxaloacetate with NADH, a broad optimum pH ranging from 8.2 to 9.4 was found with 50 mM Tris-HCl and glycine-NaOH buffers. Sodium phosphate buffer apparently activated the enzyme. The apparent Km values for oxaloacetate and NADH were 19 microM and 30 microM, respectively. The optimum pH for malate oxidation with NAD+ was 10.2 in 50 mM NaHCO3-Na2CO3 buffer. The apparent Km values for malate and NAD+ were 7.0 mM and 0.6 mM, respectively. Zinc ion, sulfite ion, p-chloromercuriphenylsulfonate and adenine nucleotides strongly inhibited the enzyme. 相似文献
16.
Factor XIII-mediated cross-linking of NH2-terminal peptide of alpha 2-plasmin inhibitor to fibrin 总被引:1,自引:0,他引:1
The NH2-terminal 12-residue peptide of alpha 2-plasmin inhibitor, Asn-Gln-Glu-Gln-Val-Ser-Pro-Leu-Thr-Gly-Leu-Lys-NH2 . AcOH, was found to be a good substrate for plasma transglutaminase (activated blood coagulation factor XIII) and rapidly incorporated into fibrin by the enzyme. A high concentration of the peptide inhibited the enzyme-mediated cross-linking of alpha 2-plasmin inhibitor to fibrin probably by competing with the inhibitor for the same site of fibrin alpha-chain. 相似文献
17.
Influence of phospholipase treatments on ligand bindings to a benzodiazepine receptor-GABA receptor-chloride ionophore complex 总被引:2,自引:0,他引:2
Treatment of rat cerebellar membranes with phospholipase A2 (PLA2) or phospholipase C (PLC) increased basal [3H]diazepam binding at 0 degrees C with concomitant disappearance of the stimulatory effect of Cl- ion on the binding. On the other hand, these treatments did not affect the stimulatory effect of GABA, nor the maximum enhancement obtained in the presence of both GABA and Cl- ion. These results suggest that PLA2 or PLC modified the phospholipids responsible for the interaction between the benzodiazepine receptor and the Cl- ionophore. This assumption was supported by the results of thermodynamic experiments which showed that the changes in thermodynamic parameters occurring after the addition of Cl- ion resembled those after PLA2 or PLC treatment. Since the effect of PLA2 was evident at very low concentrations, and a PLC concentration of at least one order of magnitude higher was required to induce a similar effect, the change of phospholipids especially to lysophospholipids seems to be of particular importance. Protein release from the membrane, which also occurs after PLA2 or PLC treatment, did not appear to be responsible for the present phenomenon. 相似文献
18.
Surface-Bound Nuclease of Staphylococcus aureus: Localization of the Enzyme 总被引:3,自引:2,他引:1
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The cellular localization of staphylococcus nuclease, previously known as an exoenzyme, was investigated, and the following results were obtained. (i) When Staphylococcus aureus cells were converted to protoplasts by cell wall lytic enzyme L-11 (a bacteriolytic enzyme purified from Flavobacterium sp. which specifically hydrolyzes amide and peptide linkages of murein layers), over 80% of the cell-bound nuclease was released into the surrounding sucrose medium. (ii) The cell-bound nuclease was associated with the cell-wall membrane fraction of mechanically disrupted cells. (iii) The nuclease activity of cell-wall membrane fractions from cells during early and late stages of protoplast formation were compared. Less activity was found in the late stage. These results suggest that nuclease may be located at or near the surface of the cells. The distribution of cell-bound nuclease in the cell-wall membrane fraction varied with the growth conditions of S. aureus. The activity of alkaline phosphatase, another surface enzyme, was also investigated. Less of this enzyme than nuclease was released when the cells were converted to protoplasts. 相似文献
19.
Surface-Bound Nuclease of Staphylococcus aureus: Purification and Properties of the Enzyme
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The surface-bound nuclease of Staphylococcus aureus liberated during formation of protoplasts was purified 1,000-fold by chromatography on phosphocellulose. Its properties were compared with those of the known extracellular nuclease, purified 200-fold by the same procedures. The adsorbance of the surface-bound nuclease on phosphocellulose was distinctly different from that of the extracellular nuclease, but other properties of the two enzymes were similar. Both enzymes had a pH optimum of about 10 and required Ca2+ for activity. Both enzymes hydrolyzed deoxyribonucleic acid (DNA) and ribonucleic acid, and denatured DNA was a better substrate than native DNA. Both enzymes were inhibited by the same metal ions. Nuclease-less mutants of S. aureus were isolated from S. aureus 209P by using N-methyl-N′-nitroso-N-nitrosoguanidine. These mutants contained neither surface-bound nor extracellular nuclease activity. These results suggest that the surface-bound and extracellular nucleases are expressed from the same cistron of S. aureus. 相似文献
20.
M Nakano K Kasai K Yoshida T Tanimoto Y Tamaki T Tobita 《Journal of biochemistry》1989,105(1):133-137
1. CD spectra showed that the fowl protamine, galline, has an unordered structure rich in reverse turns in neutral solution. Eight reverse turns were predicted to be present in the galline molecule on the basis of its amino acid sequence. Spectrophotometric analyses revealed that galline efficiently bound to DNA in 0.25 mM EDTA/10 mM Tricine-HCl, pH 7.4, but hardly so in 30 mM NaCl/3 mM sodium citrate, pH 7.0. Citrate ions bound specifically to the galline molecule, causing a conformational change in it. As a result, galline could not interact with DNA. 2. The concentration of unbound galline in a mixture of DNA and galline in 100 mM NaCl/50 mM Tricine-HCl, pH 7.4, at 37 C was determined by measurement of the intrinsic fluorescence of tyrosine residues of galline in the supernatant after ultracentrifugation of the mixture. The Scatchard plot showed positive co-operativity in the binding of galline to DNA and the binding parameters were determined: the co-operative binding constant (Kc) = 3.3 X 10(7)M-1, the co-operativity factor (q) = 800, and the number of nucleotides of DNA occupied by one galline molecule (n) = 28. The Kc and q values were intermediate between those for clupeine Z from herring sperm and S-methyl protamine from boar sperm. That is, the binding constants of protamine as to DNA decrease in the order of herring, fowl, and boar, while the co-operativities in binding increase in that order. 相似文献