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81.
Two polyclonal antibodies were raised to synthetic peptides corresponding to amino acids Ser21-Tyr35 and Lys247-Phe261 of cytochrome b561. These antibodies were used to test the native orientation of the amino and carboxyl termini of this transmembrane electron transport protein. Carboxyl-terminal epitopes were lost when intact chromaffin granules were treated with Pronase. This result indicates that the carboxyl terminus is cytoplasmically exposed and confirms a theoretical prediction obtained from hydropathy plots. Epitopes that were recognized by an amino-terminal antipeptide antibody were not removed under the same conditions. This finding implied that the amino terminus was not proteolytically accessible on the exterior of the granule. The abundance of threonine and serine residues in the amino-terminal region suggested that the amino terminus could be held in the membrane by covalent fatty acylation. Treatment of purified delipidated cytochrome b561 with hydroxylamine resulted in the release of a fatty acid hydroxamate. Sulfhydryl analysis of purified cytochrome b561 showed that all 3 cysteine residues were in the free sulfhydryl form. These observations indicate that cytochrome b561 is covalently fatty acylated and that the lipid is bound through ester linkages of serine or threonine residues. 相似文献
82.
Nasi Mian Caroline E. Anderson Andrew J. Pope Anthony R. Smith Paul S. Richardson Kim Balfre Paul W. Kent 《The Biochemical journal》1982,208(2):425-433
Chicken tracheal mucosa in vitro transported and incorporated radioactive precursors into mucins, which were secreted at a steady rate into the tracheal lumen. Secretion of mucins labelled with 35S and 3H after pulse-labelling of the mucosal layer with Na235SO4 and d-[1-3H]glucosamine as precursors was an energy-dependent process, as it was strongly inhibited by the action of respiratory-chain inhibitors, an uncoupler of oxidative phosphorylation, a metabolic blocker and a temperature shift from 41°C to 5°C. On the other hand, both cholinergic and parasympathomimetic agents considerably increased the secretion of dual-radiolabelled mucins when applied on the submucosal side of the trachea. The effect of Ca2+ was directional, since only high submucosal (3.6 or 18mm) or low luminal (zero or 0.18mm) Ca2+ massively enhanced the secretion of radiolabelled mucin compared with the mucin output measured under physiological Ca2+ conditions (1.8mm). Whereas application of ionophore A23187 on either side of the trachea significantly increased mucin output, its presence in the appropriate tracheal compartment and under appropriate Ca2+ conditions further accentuated the output of radiolabelled mucins. Addition of acetylcholine under appropriate conditions also had an additive effect on the Ca2+-stimulated secretion of mucins. Ca2+ stimulation of mucin secretion appears to be dependent on the metabolic integrity of the mucosal cells. Mucins secreted in response to high submucosal and low luminal [Ca2+] appear to consist of a number of different types of glycoproteins, as judged from their ion-exchange-chromatographic behaviour. 相似文献
83.
A. Henry Latorella Susan K. Bromberg Kent Lieber James Robinson 《Journal of phycology》1981,17(2):211-214
Fifteen nitrate assimilation-deficient mutants of the euryhaline green alga, Dunaliella tertiolecta Butcher were selected by their chlorate resistance. Ten mutants, unable to grow on NO3? but able to grow on NO2?, had no detectable nitrate reductase activity. Five mutants, unable to grow on either NO3? or NO2?, had depressed levels of both nitrate and nitrite reductase. A method for assaying methyl viologen-nitrate reductase in the presence of nitrite reductase is described. 相似文献
84.
Properties of the Photosynthetic System and DNA of Cyanophora paradoxa Cyanelles 总被引:1,自引:1,他引:0 下载免费PDF全文
The cyanelle from the photosynthetic biflagellate protist Cyanophora paradoxa has been studied in terms of its photosynthetic properties. Structurally, the cyanelle resembles unicellular cyanobacteria. The cyanelle is readily released from the host cell by means of the French press. The isolated cyanelle shows typical photosystem I and photosystem II activities as well as phenazine methosulfate-mediated photophosphorylation. The kinetic parameters Km and Vmax were determined for CO2 fixation in the cyanelle and cells of C. paradoxa and compared to a cyanobacterium. The determined values were not much different, although the cyanobacterium had a significantly greater rate of CO2 fixation, and the cyanelle was least active in this regard. Photosystem I chlorophyll-protein complex is readily isolated from the thylakoid membrane. In all these respects, the photosynthetic apparatus of the cyanelle resembles that of cyanobacteria. No nitrogen fixation activity was observed. Attempts to regenerate the isolated cyanelle were not successful, but in some cases, an unidentified cyanobacterium grew up in standing cultures of C. paradoxa cyanelles. Buoyant density data indicate that the strain of C. paradoxa we have investigated differs from that employed by others, since our strain shows a value of 1.716 grams per cubic centimeter and others report values of 1.695 and 1.691. 相似文献
85.
Simultaneous kinetic analysis of ribulose 1,5-bisphosphate carboxylase/oxygenase activities 总被引:4,自引:4,他引:0 下载免费PDF全文
An assay was developed for simultaneous kinetic analysis of the activities of the bifunctional plant enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase [EC 4.1.1.39]. [1-14C,5-3H]Ribulose 1,5-bisphosphate (RuBP) was used as the labeled substrate. Tritium enrichment of the doubly labeled 3-phosphoglycerate (3-PGA) product, common to both enzyme activities, may be used to calculate Vc/Vo ratios from the expression A/(B-A) where A and B represent the 3H/14C isotope ratios of doubly labeled RuBP and 3-PGA, and Vc and Vo represent the activities of carboxylase and oxygenase, respectively. Doubly labeled substrate was synthesized from [2-14C]glucose and [6-3H]glucose using the enzymes of the pentose phosphate pathway coupled with phosphoribulokinase. 相似文献
86.
Vidmantas A. Raisys Patrick N. Friel Patricia R. Graaff Kent E. Opheim Alan J. Wilensky 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1980,183(4)
A one-step method for extraction of diazepam, nordiazepam, and internal standard into toluene is followed by chromatographic separation and detection with either dual-wavelength high-performance liquid chromatography or electron-capture gas—liquid chromatography. Agreement between the two methods was excellent for diazepam (r = 0.99, n = 38) and good for nordiazepam (r = 0.96, n = 79) over a concentration range that included subtherapeutic, therapeutic, and toxic plasma levels. 相似文献
87.
The activity of the plasma membrane enzyme 5′-nucleotidase varies dramatically during the embryonic development of chick pectoral muscle. The specific activity is greatest at early stages of differentiation (8-day embryos), falls to a minimum on days 12–14, then rises again in older embryos. In cultured muscle cells obtained from embryonic chick muscle the 5′-nucleotidase activity is essentially absent. Muscle cells grown in the presence of bromodeoxyuridine, an inhibitor of muscle differentiation, contain enhanced levels of 5′-nucleotidase activity. These results indicate that 5′-nucleotidase may be absent in muscle fibers, but present in other cells of muscle tissue. 相似文献
88.
Estrogen receptor (ER) assays in human breast cancer tissue have proved useful in selecting patients for endocrine therapies. The absence of ER indicates hormone independent tumors and precludes the use of endocrine therapy. Patients with positive tumor ER respond to endocrine therapy at nearly twice the rate of those patients chosen by clinical criteria, although about a third of ER positive tumors in patients still do not respond. Recently, research has been directed toward increasing the accuracy of the ER assay in the ER positive group. The absolute tumor ER value and the presence of progesterone receptor appear promising in this regard. The significance of nuclear estrogen receptor is being studied. Finally, the ER status of a primary breast tumor appears to be a marker for the length of time until recurrence after mastectomy, and for survival. The ER assay may prove valuable in planning new adjuvants in the treatment of breast cancer. 相似文献
89.
Endy Chung R.Kent Rhodes Edward J. Miller 《Biochemical and biophysical research communications》1976,71(4):1167-1174
Fractionation of pepsin-solubilized collagens from several human tissues has shown that substantial quantities of collagen-like protein remain in solution under conditions leading to the precipitation of Type I, II, and III collagens. Characterization of the more soluble collagens has led to the isolation of three unique collagenous components each of which exhibit compositional features indicative of their origin from basement membranes. One of these has an apparent molecular weight of 55,000 daltons and appears to originate in endothelial basement membranes. The other two components (A chain and B chain) are somewhat larger than collagen α chains and appear to be derived from the collagen of epithelial and smooth muscle basement membranes, respectively. 相似文献
90.