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61.
P Schlesinger J S Rodman M Frey S Lang P Stahl 《Archives of biochemistry and biophysics》1976,177(2):606-614
Certain highly purified forms of rat lysosomal glycosidases, β-glucuronidase and N-acetyl-β-d-glucosaminidase, are rapidly cleared from the circulation following intravenous infusion. Several lines of evidence are presented which indicate that the primary site of enzyme uptake is the liver. Clearance of the two enzymes was unaffected by nephrectomy, whereas it was abolished by evisceration. Tissue distribution experiments with native and [125I]β-glucuronidase indicate the liver as the major, if not exclusive, site of enzyme uptake. Experiments with the isolated perfused liver showed clearance of certain enzyme preparations but not others. Those enzymes cleared by the isolated perfused liver were likewise cleared in vivo. Liver fractionation studies following infusion of large doses of β-glucuronidase revealed a rapid, short-lived increase in microsomal β-glucuronidase and a slower but larger increase in lysosomal β-glucuronidase. The results indicate that β-glucuronidase, N-acetyl-β-d-glucosaminidase, and probably other glycosidases are rapidly incorporated into the lysosomal compartment of liver. 相似文献
62.
Approximately 75% or more of the L2 and L3 juvenile stages of the free-living nematode Caenorhabditis briggsae survived freezing and thawing without loss of fertility. Optimum survival depended upon a combination of conditions: (1) pretreatment with 5% DMSO at 0 °C for 10 min, (2) 0.2 °C per minute cooling rate from 0 to ?100 °C prior to immersion into liquid nitrogen, and (3) a 27.6 °C per minute warming rate from ?196 °C to ?10 °C. Storage at ?196 °C for more than 100 days was without effect on viability or fertility. Some of the L4 (about 50%) and adult (about 3%) stages survive the routine freeze-thaw treatment. However, there was no recovery of either embryonic stages or embryonated eggs from ?196 °C under these standard conditions. Either very fast cooling (about 545 °C/min) or fast warming (about 858 °C/min) rates diminished survival of the L2 and L3 stages drastically.Scanning electron microscopy revealed that freeze-thaw survivors with aberrant swimming behavior had cuticular defects. In juvenile forms, the altered swimming motion was lost after a molt whereas as abnormal adults grew, sinusoidal movement resumed. In the L4 and adult forms the cuticular abnormalities lowered viability and fertility. It is concluded that survival of nematodes from a freeze-thaw cycle is contingent upon establishing specific cryobiological conditions by varying aspects of the procedure that gave high recoveries of L2 and L3 stages. 相似文献
63.
C B Libby W A Frey J J Villafranca S J Benkovic 《The Journal of biological chemistry》1975,250(19):7564-7573
The separate interaction of the substrate fructose 1,6-bisphosphate and a metal ion cofactor Mn2+ with neutral hexosebisphosphatase has been studied under equilibrium conditions at pH 7.5 with gel filtration and electron paramagnetic resonance measurements, respectively. Binding data for both ligands to the enzyme yielded nonlinear Scatchard plots that analyze in terms of four negatively cooperative binding sites per enzyme tetramer. Graphical estimates of the binding constants were refined by a computer searching procedure and nonlinear least squares analysis. These results are qualitatively similar to those obtained from binding studies involving teh alkaline enzyme, a modified form of hexosebisphosphatase whose pH optimum is in the alkaline pH region. Both forms of the enzyme enhance the proton relaxation rate of water protons by a factor of approximately 7 to 8 at 24 MHz, demonstrating similar metal ion environments. Teh activator Co(III)-EDTA did not affect Mn2+ binding to the neutral enzyme. In the presence of (alpha + beta)methyl-D-fructofuranoside 1,6-bisphosphate, however, two sets--each containing four Mn2+ binding sites--were observed per enzyme tetramer with loss of the negatively cooperative interaction. These results are viewed in terms of four noncatalytic and four catalytic Mn2+ binding sites. Parallel kinetic investigations were conducted on the neutral enzyme to determine specific activity as a function of Mn2+ and fructose 1,6-bisphosphate concentration. A pro-equilibrium sequential pathway model involving Mn2+-enzyme and the Mn2+-fructose 1,6-bisphosphate complex both as substrate and as an allosteric inhibitor satisfactorily fit the kinetic observations. All possible enzyme species were computed from the determined binding constants and grouped according to the number of moles of Mn2+-fructose 1,6-bisphosphate complex bound to the Mn2+-enzyme, and individual rate constants were calculated. The testing of other models and their failure to describe the kinetic observations are discussed. 相似文献
64.
Systolic time intervals are commonly used to identify changes in ventricular function. The method described facilitates measurement and calculation of many such intervals. This method utilizes a printed polygraph recording of electrocardiogram, phonocardiogram, and carotid pulse contour; a digitizing device for reading the necessary coordinates from the record; and a minicomputer for calculation of the intervals and for data analysis. Intervals related to approximately 100 pulse beats can be read, calculated, printed in chart and graph form, and subjected to some analyses in about an hour. 相似文献
65.
Native calf thymus DNA was gamma-irradiated at 500 mug/ml in 0.01 M NaCl in the presence or absence of oxygen. By irradiation, an increasing amount of DNA becomes reactive with a water-soluble carbodiimide-derivative (CMEC). In the DNA sections reactive with CMEC the nucleotide strands are separated, a phenomenon previously described as radiation-induced denaturation. The dose-effect curve for the formation of denatured DNA shows an upward-bent form; a distinct oxygen effect of about 2 is observed. By a comparative study with DNA samples, degraded partially with DNAse I, it was shown that a minor part of the radiation-induced denaturation results from the formation of the radiation-induced single strand breaks, whereas the major part is a local denaturation independent of the strand breaks. In these locally denatured regions 20 to 50 nucleotide pairs are separated. 相似文献
66.
67.
Sau-chi B. Yan Sutaeg Hwang Timothy D. Rustan William H. Frey 《Neurochemical research》1985,10(1):1-18
The soluble tubulin of human cerebral cortex, as assessed by [3H]colchicine binding of the 100,000g supernatant fraction, decreases drastically with age, 75 percent from age 0 to age 90. There is also a considerably lower concentration of high molecular weight proteins in the soluble fraction of postmortem human cerebral cortex than in that of nonhuman species. Human brain tubulin can be polymerized into microtubules with DEAE-dextran. The DEAE-dextran induced microtubules are stable to cold temperature (4°) and calcium. However, in the presence of 1 M glutamate, the microtubules become cold labile and depolymerize at 4°. Thus we have developed a novel method for purifying polymerization competent tubulin from fresh or frozen human cerebral cortex. Human brain tubulin purified by our novel method is very similar to tubulin from the brains of other mammals in molecular weight, amino acid composition, polymerization-depolymerization parameters, and structural dimensions of the microtubules formed.Some aspects of this work have been published as an abstract in 1981. Fed. Proc. 40:1548. 相似文献
68.
Anna Kletzmayr Flurina Clement Frey Miriam Zimmermann Daniel Eberli Christopher Millan 《Biotechnology journal》2020,15(5)
The microenvironment plays a major role in conferring chemoresistance to cancer cells. In order to better inform clinical response to chemoresistance, preclinical models that recapitulate its hallmark features are needed to enable screening for resistance‐specific therapeutic targets. A novel platform for seeding cancer cells in 3D hydrogels is presented utilizing derivatives of chitosan and alginate that, critically, is amenable to high throughput screening: cell seeding in hydrogels, media changes, dosing of anticancer compounds, and cell viability assays are all automated using a standard and commercially available liquid handling robot. Culture in these hydrogels elicits resistance in ovarian, lung, and prostate cancer cells to treatment by doxorubicin and paclitaxel. In correlation, proteomics analysis of SKOV3 cells cultured in 3D reveals enrichment of proteins associated with extreme drug resistance including HMOX1 and ALDH2. Subsequently, therapeutic antibodies targeted to tumor‐associated antigens upregulated in 3D cultures are shown to have higher efficacy compared to 2D cultures. Collectively, this automated 3D cell culture platform provides a powerful tool with utility in identification of drugs that may overcome chemoresistance. 相似文献
69.
Ilse Storch Johannes Penner Thomas Asbeck Marco Basile Jürgen Bauhus Veronika Braunisch Carsten F. Dormann Julian Frey Stefanie Grtner Marc Hanewinkel Barbara Koch Alexandra‐Maria Klein Thomas Kuss Michael Pregernig Patrick Pyttel Albert Reif Michael Scherer‐Lorenzen Gernot Segelbacher Ulrich Schraml Michael Staab Georg Winkel Rasoul Yousefpour 《Ecology and evolution》2020,10(3):1489-1509
Retention forestry, which retains a portion of the original stand at the time of harvesting to maintain continuity of structural and compositional diversity, has been originally developed to mitigate the impacts of clear‐cutting. Retention of habitat trees and deadwood has since become common practice also in continuous‐cover forests of Central Europe. While the use of retention in these forests is plausible, the evidence base for its application is lacking, trade‐offs have not been quantified, it is not clear what support it receives from forest owners and other stakeholders and how it is best integrated into forest management practices. The Research Training Group ConFoBi (Conservation of Forest Biodiversity in Multiple‐use Landscapes of Central Europe) focusses on the effectiveness of retention forestry, combining ecological studies on forest biodiversity with social and economic studies of biodiversity conservation across multiple spatial scales. The aim of ConFoBi is to assess whether and how structural retention measures are appropriate for the conservation of forest biodiversity in uneven‐aged and selectively harvested continuous‐cover forests of temperate Europe. The study design is based on a pool of 135 plots (1 ha) distributed along gradients of forest connectivity and structure. The main objectives are (a) to investigate the effects of structural elements and landscape context on multiple taxa, including different trophic and functional groups, to evaluate the effectiveness of retention practices for biodiversity conservation; (b) to analyze how forest biodiversity conservation is perceived and practiced, and what costs and benefits it creates; and (c) to identify how biodiversity conservation can be effectively integrated in multi‐functional forest management. ConFoBi will quantify retention levels required across the landscape, as well as the socio‐economic prerequisites for their implementation by forest owners and managers. ConFoBi's research results will provide an evidence base for integrating biodiversity conservation into forest management in temperate forests. 相似文献
70.