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121.
The role of site-specific N-glycosylation in secretion of soluble forms of rabies virus glycoprotein 总被引:1,自引:1,他引:0
Wojczyk BS; Stwora-Wojczyk M; Shakin-Eshleman S; Wunner WH; Spitalnik SL 《Glycobiology》1998,8(2):121-130
Rabies virus glycoprotein is important in the biology and pathogenesis of
neurotropic rabies virus infection. This transmembrane glycoprotein is the
only viral protein on the surface of virus particles, is the viral
attachment protein that facilitates virus uptake by the infected cell, and
is the target of the host humoral immune response to infection. The
extracellular domain of this glycoprotein has N- glycosylation sequons at
Asn37, Asn247, and Asn319. Appropriate glycosylation of these sequons is
important in the expression of the glycoprotein. Soluble forms of rabies
virus glycoprotein were constructed by insertion of a stop codon just
external to the transmembrane domain. Using site-directed mutagenesis and
expression in transfected eukaryotic cells, it was possible to compare the
effects of site-specific glycosylation on the cell-surface expression and
secretion of transmembrane and soluble forms, respectively, of the same
glycoprotein. These studies yielded the surprising finding that although
any of the three sequons permitted cell surface expression of full-length
rabies virus glycoprotein, only the N-glycan at Asn319 permitted secretion
of soluble rabies virus glycoprotein. Despite its biological and medical
importance, it has not yet been possible to determine the crystal structure
of the full-length transmembrane form of rabies virus glycoprotein which
contains heterogeneous oligosaccharides. The current studies demonstrate
that a soluble form of rabies virus glycoprotein containing only one sequon
at Asn319 is efficiently secreted in the presence of the N-glycan
processing inhibitor 1-deoxymannojirimycin. Thus, it is possible to purify
a conformationally relevant form of rabies virus glycoprotein that contains
only one N-glycan with a substantial reduction in its microheterogeneity.
This form of the glycoprotein may be particularly useful for future studies
aimed at elucidating the three-dimensional structure of this important
glycoprotein.
相似文献
122.
Duplication of the atrioventricular (AV) valves is a rare anomaly, more commonly seen in the mitral position. Only 13 cases have been reported in the literature.(1-10) The case described here represents an unusual variant of this anomaly and provides an opportunity to review the pathology and embryologic genesis of duplication deformities of the AV valves. 相似文献
123.
During five consecutive daily test sessions, 10 men and women rated the relative intensity and pleasantness of breath odors from 14 males and 19 females on a no-oral-hygiene regimen. In addition, the likely gender of the donor of each odor was also estimated. The breath odors of males were rated, on the average, as more intense and less pleasant than the breath odors of females. Women consistently gave lower pleasantness ratings to the odors than did men. Both the male and female judges assigned the breath odors to the correct gender classes at a frequency unlikely due to chance, although the females were more accurate in this regard. An inverse relation between breath odor intensity and pleasantness was noted. Systematic changes in the rated intensity and pleasantness of the odors were present across the 5 days of the study period. These data suggest that differences exist between the breath odors of men and women, and that humans, like many other mammals, may be capable of assessing gender from oral odors. However, such assignments conceivably reflect the strategy of assigning stronger and less pleasant odors to the male category, and weaker and less unpleasant odors to the female category, regardless of the true sex of the odor donor. 相似文献
124.
125.
M Schlame S Shanske S Doty T K?nig T Sculco S DiMauro T J Blanck 《Journal of lipid research》1999,40(9):1585-1592
Cardiolipin is a specific mitochondrial phospholipid that is present in mammalian tissues in low concentration. To measure cardiolipin in small biopsies from patients with mitochondrial disease, we developed a new technique that can detect subnanomolar levels of well-resolved molecular species, the most abundant of which are tetralinoleoyl-cardiolipin (L(4)) and trilinoleoyl-oleoyl-cardiolipin (L(3)O). To this end, a fluorescence-labeled derivative of cardiolipin (2-[naphthyl-1'-acetyl]-cardiolipin dimethyl ester) was formed and analyzed by high performance liquid chromatography. Cardiolipin was measured in skeletal muscle biopsies from 8 patients with mitochondrial disease and in 17 control subjects. In 5 patients with mitochondrial disease, cardiolipin content was higher than normal (2. 4;-7.0 vs. 0.4;-2.2 nmol/mg protein). In 3 patients with mitochondrial disease, the L(4)/L(3)O ratio was lower than normal (2;-4 vs. 4;-6). Cardiolipin was also measured in various rat and dog muscle tissues. The L(4)/L(3)O ratio was higher in condensed "muscle" type mitochondria (heart ventricle, skeletal muscle, ratios 4;-7) than in orthodox "liver" type mitochondria (liver, smooth muscle, heart auricular appendage, H9c2 myoblasts, ratios 0.4;-3), suggesting that the L(4)/L(3)O proportion is important for cristae membrane structure. We concluded that the L(4)/L(3)O ratio is a tissue-specific variable that may change in the presence of mitochondrial disease. The new method is suitable to measure cardiolipin in muscle biopsies in order to estimate concentration of mitochondria. 相似文献
126.
Recently a novel measure of olfactory function, the Sniff Magnitude Test (SMT), was developed that relies on changes in inhalation in response to an odor. The relationship of this unique test to that of other olfactory tests has received little investigation. In this study, we assessed, in 132 patients presenting to a chemosensory disorders clinic, the relationship of SMT scores to those from 3 standardized psychophysical tests: the University of Pennsylvania Smell Identification Test (UPSIT), a phenyl ethyl alcohol odor detection threshold test, and a short-term odor memory/discrimination test. SMT scores were roughly related to olfactory dysfunction categories defined for the UPSIT and correlated moderately with the other tests. Malodors (1% and 3% methylthiobutyrate [MTB], 1% ethyl 3-mercaptoproprionate) exhibited stronger correlations than nonmalodors (3% phenyl ethyl alcohol [PEA], 3% amyl acetate, 3% n-butanol) and elicited greater sniff suppression. In a principal component analysis, the SMT measures loaded on components different from those of the other tests, which loaded on a separate component. Anticipatory responses (i.e., smaller sniffs) occurred across trials for the first malodor (1% MTB), but not for the first nonmalodor (3% PEA), that was encountered. These results, along with those of an earlier factor analysis, suggest that sniff magnitude is influenced by odorant quality and intensity, as well as by cognitive factors. 相似文献
127.
Pourmand EP Binderman I Doty SB Kudryashov V Boskey AL 《Journal of cellular biochemistry》2007,100(1):43-57
The calcification of cartilage is an essential step in the process of normal bone growth through endochondral ossification. Chondrocyte apoptosis is generally observed prior to the transition of calcified cartilage to bone. There are, however, contradictory reports in the literature as to whether chondrocyte apoptosis is a precursor to cartilage calcification, a co-event, or occurs after calcification. The purpose of this study was to test the hypothesis that chondrocyte apoptosis is not a requirement for initial calcification using a cell culture system that mimics endochondral ossification. Mesenchymal stem cells harvested from Stages 21-23 chick limb buds were plated as micro-mass cultures in the presence of 4 mM inorganic phosphate (mineralizing conditions). The cultures were treated with either an apoptosis inhibitor or stimulator and compared to un-treated controls before the start of calcification on day 7. Inhibition of apoptosis with the caspase inhibitor Z-Val-Ala-Asp (O-Me)-fluoromethylketone (Z-VAD-fmk) caused no decreases in calcification as indicated by radioactive calcium uptake or Fourier transform infrared (FT-IR) analysis of mineral properties. When apoptosis was inhibited, the cultures showed more robust histological features (including more intense staining for proteoglycans, and more intact cells within the nodules as well as along the periphery of the cells as compared to untreated controls), more proliferation as noted by bromo-deoxyuridine (BrdU) labeling, decreases in terminal deoxynucleotidyl transferase (Tdt)-mediated dUTP nick-end labeling (TUNEL) staining, and fewer apoptotic bodies in electron microscopy. Stimulation of apoptosis with 40-120 nM staurosporine prior to the onset of calcification resulted in inhibition of calcium accretion, with the extent of total calcium uptake significantly decreased, the amount of matrix deposition impaired, and the formation of abnormal mineral crystals. These results indicate that chondrocyte apoptosis is not a pre-requisite for calcification in this culture system. 相似文献
128.
Despite the recent introduction of biological response modifiers and potent new small-molecule antirheumatic drugs, the efficacy of methotrexate is nearly unsurpassed in the treatment of inflammatory arthritis. Although methotrexate was first introduced as an antiproliferative agent that inhibits the synthesis of purines and pyrimidines for the therapy of malignancies, it is now clear that many of the anti-inflammatory effects of methotrexate are mediated by adenosine. This nucleoside, acting at one or more of its receptors, is a potent endogenous anti-inflammatory mediator. In confirmation of this mechanism of action, recent studies in both animals and patients suggest that adenosine-receptor antagonists, among which is caffeine, reverse or prevent the anti-inflammatory effects of methotrexate. 相似文献
129.
Corn samples were collected in 1999 from three departments of Entre Réos province, Argentina, and were surveyed for mould
contamination and natural occurrence ofFusarium mycotoxins, ochratoxin A and aflatoxins.Fusarium verticillioides was the most prevalent fungal species recorded at all departments. Zearalenone, deoxynivalenol and ochratoxin A were not
found in any samples. Only one of the 52 corn samples analysed was contaminated with aflatoxin B1 (17 μg/kg). Fumonisin B1 was found in 58 % of samples (range of positive samples: 47– 3,347 μg/kg), fumonisin B2 in 33.0 % (range of positive samples: 23–537 μg/kg) and fumonisin B3 in 25.0 % (range of positive samples: 24–287 μg/kg) of them. This is the first report on the natural occurrence of mycotoxins
in corn from Entre Ríos province, Argentina. Levels of fumonisins were lower than detected in other Argentinian provinces. 相似文献
130.
Detection of endogenous alkaline phosphatase activity in intact cells by flow cytometry using the fluorogenic ELF-97 phosphatase substrate. 总被引:1,自引:0,他引:1
BACKGROUND: The alkaline phosphatase (AP) substrate 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-(3H)-quinazolinone (ELF((R))-97 for enzyme-labeled fluorescence) has been found useful for the histochemical detection of endogenous AP activity and AP-tagged proteins and oligonucleotide probes. In this study, we evaluated its effectiveness at detecting endogenous AP activity by flow cytometry. METHODS: The ELF-97 phosphatase substrate was used to detect endogenous AP activity in UMR-106 rat osteosarcoma cells and primary cultures of chick chondrocytes. Cells were labeled with the ELF-97 reagent and analyzed by flow cytometry using an argon ultraviolet (UV) laser. For comparison purposes, cells were also assayed for AP using a Fast Red Violet LB azo dye assay previously described for use in detecting AP activity by flow cytometry. RESULTS: The ELF-97 phosphatase substrate effectively detected endogenous AP activity in UMR-106 cells, with over 95% of the resulting fluorescent signal resulting from AP-specific activity (as determined by levamisole inhibition of AP activity). In contrast, less than 70% of the fluorescent signal from the Fast Red Violet LB (FRV) assay was AP-dependent, reflecting the high intrinsic fluorescence of the unreacted components. The ELF-97 phosphatase assay was also able to detect very low AP activity in chick chondrocytes that was undetectable by the azo dye method. CONCLUSIONS: The ELF-97 phosphatase assay was able to detect endogenous AP activity in fixed mammalian and avian cells by flow cytometry with superior sensitivity to previously described assays. This work also shows the applicability of ELF-97 to flow cytometry, supplementing its previously demonstrated histochemical applications. 相似文献