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51.
K Inada S Endo K Takahashi M Suzuki T Narita T Yoshida H Suda T Komuro M Yoshida 《Microbiology and immunology》1991,35(4):303-314
We established a new method of plasma treatment for the removal of interfering factors in the plasma to allow detection of endotoxin by limulus test. The limulus test used was an endotoxin-specific chromogenic test, the Endospecy test. Perchloric acid (PCA) treatment and centrifugation (PCA method) is usually used to remove interfering factors from plasma, with the precipitate being discarded and the supernatant used to detect endotoxin. As the solubilized precipitates of endotoxin-spiked plasma and some patient plasma were found to contain the Endospecy activity, we have devised a new method assaying endotoxin in both the supernatant and precipitate. This study confirmed that the solubilized precipitate of endotoxin-spiked plasma had Endospecy activity and found that the precipitate had other endotoxin activities, such as lethality in galactosamine-sensitized mice and pyrogenicity in rabbits. We also confirmed that interfering factors were completely removed from plasma samples by this new method. The endotoxin level after the new PCA method was found to be about 8 times higher than that determined after PCA treatment and the new PCA method surpasses the conventional PCA method with regard to the positive rate of endotoxin contents in clinical samples. These results indicate that the new PCA method is superior to the PCA method as a plasma pretreatment method for limulus test. 相似文献
52.
Sensitive detection of viral antigens with a new method, "laser magnet immunoassay". 总被引:1,自引:0,他引:1
H Mizutani M Suzuki H Mizutani K Fujiwara S Shibata K Arishima M Hoshino H Ushijima H Honma T Kitamura 《Microbiology and immunology》1991,35(9):717-727
A new method, "laser magnet immunoassay" (LMIA), has been developed for sensitive detection of viral antigens. Target viruses captured on microbeads were made to react with antibodies labeled with magnetite particles. In a magnetic field, magnetically labeled antigens dispersed in water were attracted to and concentrated at one point on the surface, resulting in the lifting up of a small surface area. A laser beam which was incident on the point reflected, making an interference fringe. The intensity of the fringe indicates the amount of the magnetite conjugated with antigen. A very low concentration of antigens, such as 5 particles of influenza virus and 0.1 pg/ml of human immunodeficiency virus (HIV) p24 antigen in human serum, could be detected by this method. Application of this method to diagnoses of viral diseases in early stages is discussed. 相似文献
53.
Gene organization in the region containing a new gene involved in chromosome partition in Escherichia coli. 总被引:21,自引:9,他引:12 下载免费PDF全文
A new mutation, parC, causing abnormal chromosome segregation was identified in two thermosensitive mutants of Escherichia coli. The thermosensitive growth of the mutants was corrected by pLC4-14 in the Clarke-Carbon collection. This plasmid carries a putative gene which can suppress the cell division defect due to ftsI (pbpB) and has hence been termed sufI (sui). The nearness of parC to metC was confirmed, and cotransduction frequency of parC was 59% with metC and 20% with glc. The parC-sufI region was analyzed by subcloning the chromosome region of pLC4-14. The parC and the sufI gene products were electrophoretically identified as proteins of 75 and 55 kilodaltons (kDa), respectively. The allelism of parC+ on pLC4-14 to parC1215 was confirmed by cloning parC1215. The sufI gene appeared to be dispensable for cell viability, and overproduction of its product caused suppression of ftsI. An essential gene coding for a 25-kDa protein was found between the parC and the sufI gene. These three genes were transcribed in the same direction and may be organized into an operon, with parC to the proximal side and with internal promoters at least for the distal genes. The localization of the gene products was examined in maxicells. The sufI protein was synthesized as a precursor which could be chased into a mature form. The major part of the mature form was found in the soluble fraction. The 25-kDa protein was found almost exclusively in the membrane fraction. The parC protein was associated with the membrane fraction in the presence of Mg2+ but found in the soluble fraction when Mg2+ was sequestered with EDTA. 相似文献
54.
Gene structure of human thrombomodulin, a cofactor for thrombin-catalyzed activation of protein C 总被引:2,自引:0,他引:2
T Shirai S Shiojiri H Ito S Yamamoto H Kusumoto Y Deyashiki I Maruyama K Suzuki 《Journal of biochemistry》1988,103(2):281-285
The gene coding for human thrombomodulin, a thrombin receptor on endothelial cells and a cofactor for the activation of anticoagulant protein C zymogen, was isolated from a human genomic library by employing human thrombomodulin cDNA as a probe. The nucleotide sequences of the gene and the adjacent 5' and 3' flanking regions were then determined. The nucleotide sequence of this gene with approximately 3.7 kilobase pairs was identical to that of the cDNA, indicating that the gene for human thrombomodulin is free of introns. Hybridization data showed that there is only a single thrombomodulin gene in the human genome. 相似文献
55.
The transport of 1,5-anhydro-D-glucitol (AG) across plasma membranes was investigated in rat hepatoma cells, Reuber H-35. The AG uptake by the cells showed a concentration gradient dependency: the uptake was saturated within 40 s, which was less than one-third of the saturation time for 2-deoxy-D-glucose (DG) uptake. Furthermore, the Km value of the transport system for AG was higher than 100 mM. Though AG has a pyranoid structure resembling that of glucose, AG did not compete for cellular uptake with DG, D-glucose or 3-O-methyl-D-glucose, which are taken into cells through the glucose transporters. Conversely, the DG transport was not inhibited by AG at concentrations up to 50 mM. AG transport was hardly inhibited by 10 microM cytochalasin B, which strongly inhibits glucose transporters. In contrast, the AG transport was inhibited by 100 microM phloretin much more strongly than the DG transport when cells were preincubated with the inhibitor; the inhibition constant was 28.0 microM. The AG transport was not inhibited by 100 microM phloridzin, while the DG uptake was slightly inhibited by phloridzin. On the basis of these observations we propose that the AG uptake into rat hepatoma cells is mediated by a carrier distinct from glucose transporters. 相似文献
56.
Y Kawamura-Konishi H Suzuki 《Biochemical and biophysical research communications》1988,156(1):348-354
We prepared normal and modified alpha and beta globulin chains in which C-terminal residues were enzymatically removed. The CD spectra of the deoxy form of these chains and the reconstituted modified Hb's were measured in the Soret region. The CD spectra of the modified Hb's were markedly different from the arithmetic means of respective spectra of their constituent chains. This difference was ascribed to the interaction between alpha 1 and beta 1 subunits to make the alpha 1 beta 1 dimer. The peak wavelength of the difference CD spectra could be classified into two groups, one was 433 +/- 1 nm and the other 437 +/- 1 nm. A comparison of this classification with the previously identified quaternary structures revealed that the R and T structures showed a maximum of the difference CD spectra at 437 +/- 1 nm and 433 +/- 1 nm, respectively. These results indicated that the R and T structures differed in the interaction between alpha 1 and beta 1 subunits. 相似文献
57.
M Suzuki Y Kawaguchi M Momose T Morita K Yokoyama T Miyahara 《Biochemical and biophysical research communications》1988,150(3):1193-1198
We examined the effects of several forms of vitamin D added to renal brush-border membrane suspensions on phosphate and glucose transport and on membrane fluidity. The 1,25-D stimulated and the other vitamin D decreased phosphate uptake. In contrast, glucose uptake was not affected by the treatment of vitamin D. The 1,25-D resulted in a significant shift of the lower transition temperature in Arrhenius plots for phosphate, but not for glucose uptakes, from 15 degrees C to 11.5 degrees C. These data indicate that the 1,25-D may alter membrane fluidity, limited to the phosphate transporter, thus affecting the phosphate uptake. 相似文献
58.
The purpose of this study was to examine the development of newly born C57BL/KsJ-dbm mice produced from eggs fertilized in vitro. The embryos derived from fertilization in vitro (which was performed by using db/db eggs and adrenalectomized db/db (Adrex) spermatozoa,) were transferred to the oviduct of MRL/MpJ pseudopregnant recipients 30 hr after insemination. 376 of these embryos yielded 65 young. Weight gain and urine glucose, plasma glucose and insulin levels were measured in these young as well as in Adrex males. The young produced by fertilization in vitro showed hyperglycemia, hyperinsulinemia and obesity. The physiological abnormalities in these young were similar to those in db/db young produced by natural mating between heterozygote (db/+) males and females. Adrex males did not show hyperglycemia but did show hyperinsulinemia. These results indicate that in vitro fertilization and embryo transfer is an effective means of producing fetuses or newborns with an overt genotype in genetically diabetic obese (db) mice. 相似文献
59.
The analbuminemic rat strain established by Nagase et al. (Nagase, S., Shimamune, K., and Shumiya, S. (1979) Science 205, 590-591) exhibits hereditary deficiency in albumin biosynthesis. Serum bilirubin concentration is rather lower in homozygous (aa) rats (0.009 +/- 0.002 mg/dl) as compared with heterozygous (Aa) rats (0.047 +/- 0.009 mg/dl) or wild-type Sprague-Dawley (AA) rats (0.034 +/- 0.006 mg/dl) as evidenced by high pressure liquid chromatography analysis of bilirubin. After intravenous administration of various amounts of [heme-3H]hemoglobin in rats, [3H]bilirubin derived from [3H]heme of hemoglobin in vivo is more efficiently excreted into bile in aa rats than in Aa or AA rats. [3H]Bilirubin is exclusively bound with high-density lipoprotein (HDL) in aa rats, and a significant amount of [3H]bilirubin is shown to bind with HDL in Aa or AA rats in vivo. Scatchard plots revealed that [3H]bilirubin is bound with HDL in three binding modes depending on the molar ratio of [3H]bilirubin to HDL: Kd = 0.8 X 10(-7) M (molar ratio, 0.02-0.06), Kd = 1.6 X 10(-6) M (molar ratio, 0.06-0.41), and Kd = 1.2 X 10(-4) M (molar ratio, 0.79-9.02). Even under extreme conditions of excess hemoglobin administration, the molar ratio remains under 0.041; and thus, expected the Kd value would remain around 0.8 X 10(-7) M. Binding of [3H]bilirubin to rat serum albumin revealed two distinct binding modes depending on the molar ratio of [3H]bilirubin to rat serum albumin: Kd = 3.6 X 10(-7) M (molar ratio, 0.03-0.21), and Kd = 5.0 X 10(-6) M (molar ratio, 0.21-2.46). Under physiological conditions in Aa or AA rats, the former mode would be more reliable than the latter. Thus, HDL could bind with approximately 4.5 times higher affinity than rat serum albumin in Aa or AA rats under physiological conditions in vivo. 相似文献
60.
Microtubule-binding domain of tau proteins 总被引:12,自引:0,他引:12
H Aizawa H Kawasaki H Murofushi S Kotani K Suzuki H Sakai 《The Journal of biological chemistry》1988,263(16):7703-7707
Limited chymotryptic digestion of whole tau proteins produced a fragment of Mr 14,000 (CT14), which was able to bind to microtubules reconstituted from tubulin alone in the presence of taxol. This fragment was also found to persist in microtubules when microtubules consisting of tau proteins and tubulin were digested by chymotrypsin. Analysis of amino acid composition revealed that CT14 was rich in lysine and proline residues, suggesting unique structure of microtubule-binding domain of tau proteins. Amino-terminal sequence of CT14 was determined to be Ser-Ser-Pro-Gly-Ser-Pro-Gly-Thr-Pro-Gly-Ser-Arg-Ser-Arg-X-Pro-Ser-Leu-Pr o. No heterogeneity was detected in this amino-terminal sequence of 19 residues. Five species of polypeptides consisting of tau proteins were separated from each other by gel electrophoresis and subjected to chymotryptic digestion. CT14 was produced from each of the tau polypeptides by chymotryptic digestion, indicating that all tau polypeptides have a common microtubule-binding domain. 相似文献